Staples torrance ca: Staples 22025 Hawthorne Blvd, Torrance, CA 90503

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Staples Locations in California Torrance

You are on the page of Staples California Torrance where all the information is available about the contact, phone, addresses and services.

In this store of Staples you can find out the price range of the all products which you can see online or in-store.

Address :

22025 Hawthorne Blvd, Torrance, California, USA

Postal Code : 90503
Opening Hours :

Monday – Friday: 8:00 am – 9:00 pm Saturday: 9:00 am – 9:00 pm Sunday: 10:00 am – 6:00 pm

Phone : 3105403093
Fax : 3105403974

All stores might not offer the same variety but they mostly have the typical range of products that would be available at any store of Staples.

Staples is among the biggest brands that retail best electronics, stationery and office supplies in the whole country.

Since, I advice you to have a rough look at the product range before you go shopping for electronics or professional office supplies.

Staples has also savings on product ranges randomly and anytime we can come across with this type offers on their website.

We’ll also focus on these deals and coupons you might print or use on the internet sites.

From technology range of Staples you can see:

Apple; contains offers of iPad, iPod and iPhone accessories in general and you can see products related to Mac accessories.

Cell Phones; you can see price range for smart phones, accessory range suitable with them.

Many more aisle of Staples which you can find at Staples Locations California Torrance are actually available in the range of products.

Staples California Torrance Features

  • Mobile Phones
  • Full-service UPS® Shipping
  • Buy online.Pickup in store
  • Technology Services
  • Computer Workstation
  • Ship to Store
  • Copy & Print Services
  • UPS® Prepaid Drop-off
  • Mobile Printing

Home > Staples Store Locator > California > Torrance

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Staples – Torrance, CA 90505

So much more than office and school supplies! Find a growing product selection including cleaning and safety supplies, furniture, iPads and Apple products, technology, paper, ink, electronics, computer repair, copy & printing services, phone repair services, and water damaged phone repair services.

Chain retailer offering brand-name electronics & print services, plus office supplies & furniture.

Located In: Airport Plaza Shopping Center

Service Options:  In-store shopping · Curbside pickup · Delivery

Health and Safety:  Mask required · Staff wear masks · 

Hours

Friday8AM–9PM
Saturday9AM–9PM
Sunday10AM–6PM
Monday8AM–9PM
Tuesday8AM–9PM
Wednesday8AM–9PM
Thursday8AM–9PM

Address and Contact Information

Address: 2748 Pacific Coast Hwy, Torrance, CA 90505 Phone: (310) 784-2410 Website: https://stores.staples.com/ca/torrance/2748-pacific-coast-highway?cmpid=st_ll_goo&cid=LS:RETAIL:G

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Related Web Results

Staples® Torrance, CA, Torrance, 90503 | Store Details

Visit this Torrance Staples® for office supplies & furniture, school supplies, printers, ink & toner, copy paper, computers, technology, and more.




Review Summary

D. Moe

This location has a good parking lot which affords easy access to store.

James Steininger

The staff were inexperienced in using standard Windows print settings options.

Peter Balner

Old shabby store with poor inventory and selection, but sky high prices.

Jasmine Robello (Jazz)

Customer service is great here! Employees are always helpful and kind. Wish I can remember all their names but Erin goes above and beyond for the customers. Thanks for making my shopping experience the best. 🙂

C

Best Place For Office Things And Electronics.

thebergytv

Nice cashier… Didn’t ring me up properly… Price matched 100% instead of 110% with best buy… I am not going to contest in person during Corona… Would rather handle with corporate in the safety of my own home…

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Staples 2748 Pacific Coast Hwy, Torrance, CA 90505

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Office Equipment & Supplies, Autographic Registers & Supplies, Computer & Equipment Dealers

COUPONS

  • PHOTOS AND VIDEOSDETAILS

    THAT WAS EASY.

    General Info
    With a focus on the community of small businesses and consumers, Staples Connect provides innovative products, services and inspiration essential to the new ways of working and learning. Staples Stores and Staples Connect retail locations serve millions of customers from entrepreneurs and small businesses to remote workers, parents, teachers, and students. Explore Staples Connect at a local Staples Store or online at StaplesConnect.com.
    Hours
    Regular Hours
    Mon – Fri:
    Sat:
    Sun:
    Categories
    Office Equipment & Supplies, Autographic Registers & Supplies, Computer & Equipment Dealers, Computer Printers & Supplies, Copying & Duplicating Service, Office Furniture & Equipment, Printing Services
    Payment Options
    Brands
    3M, Amazon, Apple, Avery, BIC, Brother, Canon, Clorox, Crayola, Dell, Epson, Expo, HP, Intuit, Logitech, Microsoft, Moleskine, Netgear, Paper Mate, Pentel, Perk, Pilot, Post It, Scotch, Seagate, Sharpie, Swingline, TruRed, Zagg
    Neighborhoods
    Torrance, Walteria
    Other Link

    http://www. staples.com

    Other Information

    Parking: Lot, Private

    Bike Parking: No

    Price Range : Average

    Specialties: Software, Chairs, Cabinetry, Bookcases, Mice, Desks, Custom Work, Boxes, Fax Machines, Batteries, Computer Paper, Tables, Cabinets, Armoires, Bookshelves, Toner, Printers, Printer Ink, Office Supplies, Office Furniture, Computers, Back To School Supplies

    Provided by

    Data provided by one or more of the following: DexYP, Data Axle, Yext.

    Walmart Office Supply Store in Torrance, CA | Office Phones, Paper, Calculators | Serving Southwood

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    We’re working hard to get back up & running.

    • Temporarily on hold, due to high demand.
    • Please keep checking back for availability.
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    Tell us what you need

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    How to get to Staples in Torrance by Bus

    Public Transportation to Staples in Torrance

    Wondering how to get to Staples in Torrance, United States? Moovit helps you find the best way to get to Staples with step-by-step directions from the nearest public transit station.

    Moovit provides free maps and live directions to help you navigate through your city. View schedules, routes, timetables, and find out how long does it take to get to Staples in real time.

    Looking for the nearest stop or station to Staples? Check out this list of stops closest to your destination: Pacific Coast Hwy at Crenshaw Bl; Pacific Coast Hwy + Crenshaw Bl; Pacific Coast Hwy & Crenshaw Blvd (Westbound).

    You can get to Staples by Bus. These are the lines and routes that have stops nearby – Bus: 232

    Want to see if there’s another route that gets you there at an earlier time? Moovit helps you find alternative routes or times. Get directions from and directions to Staples easily from the Moovit App or Website.

    We make riding to Staples easy, which is why over 930 million users, including users in Torrance, trust Moovit as the best app for public transit. You don’t need to download an individual bus app or train app, Moovit is your all-in-one transit app that helps you find the best bus time or train time available.

    For information on prices of Bus, costs and ride fares to Staples, please check the Moovit app.

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    Big John going, goning. going…gone!Big John going. going. going….gone

    90,000 Pelican. Professional Cases Dave Parker, founder and president of Pelican, has been fond of scuba diving since his youth. To see new underwater landscapes, he traveled to the most exotic parts of the world. Gaining invaluable experience of “underwater life”, Parker pondered a lot about how to improve the devices for his favorite hobby.

    He strove to help his like-minded people get even more pleasure from underwater travel.It was for this purpose that Dave Parker created his company in 1975.

    The company’s first patented product was the Pelican Float.

    This invention of Dave was quite simple – however, as everything is ingenious! The float and string were painted a bright yellow color, clearly visible under water, even at great distances. Such floats made it much easier for divers to re-find the target and navigate underwater.

    The main associate and the first co-founder of Dave was his wife Arline, an airline stewardess.Together they worked to create and design their products. Dave designed the equipment himself and installed the assembly line required for production. Arline handled packaging, shipping and accounting of finished products in her spare time.

    Scuba diving enthusiasts began to purchase products. They learned about Dave’s invention from advertisements posted in specialized diving equipment stores. Soon, in order to expand the company, Dave brought in an assistant – Jeri Lineham.Jerry, who graduated from the University of Southern California Business School, took over the financial side of the venture.

    Gradually from a small production facility in a garage, the company developed into an international marketing and manufacturing company. All technology for the production of cases and flashlights is patented and certified. The company has a series of unique injection molding machines and modern production facilities. Today it employs more than 500 people, and the production area is about 20 thousand square meters. “Pelican products” is constantly expanding its range of products, finding more and more new areas for its application.The company carries out regular supplies to more than 60 countries around the world. All products are manufactured in the USA, Torrance, California.

    Every year the company launches new models of cases and flashlights on the market. Today the assortment of “Pelican Products” includes 25 models of protective cases and 35 models of professional flashlights. Only one thing has been and remains unchanged – this is the guarantee of the excellent quality of its products.

    For 30 years, Peli cases have received well-deserved recognition all over the world. Peli cases are ranked 1st in the category of protective cases from all known and produced to date by other brands.We are glad to offer you more than 30 unique models of protective cases. The design and material from which they are made are protected by copyright patents.

    Peli cases have found application in all areas of human activity: in industry and science, in the army and navy, in fire and rescue operations, in medicine and sports, mountain travel and diving.

    Pelican provides a unique lifetime warranty on its cases.

    Robust case design
    Patented high pressure injection molding technology creates extra strong and durable plastic for cases.
    The copolymer polypropylene used in the Peli protective case consists of a plastic mesh core and solid walls, which is much stronger than a solid solid wall structure. The case body is capable of withstanding high shock and vibration loads. The cases have certificates of conformity of military and civilian departments of many countries of the world.

    Strong Latch Lock
    Introducing the new Latch Lock. This is the classic design of the clamping bracket, complemented by a more rational application of the laws of physics: the auxiliary movement, which acts as a lever during the initial stage of the release of the latch, allows it to be opened with a slight push.There are no springs or metal parts in the construction of the lock. Absolute guarantee for the operation of the latch lock. The lock is made of ABS plastic.

    Automatic atmospheric valve
    New design of the automatic atmospheric valve. The inner membrane made of Teflon, equalizes the external atmospheric pressure and the pressure inside the case, and does not allow moisture to penetrate into the case. Equalization of pressure in the case is necessary when the altitude or ambient temperature changes.

    Waterproof o-ring
    The secret of the waterproof case: tongue-and-groove connection using a neoprene o-ring to ensure water tightness. Full tightness is guaranteed at a depth of 10 meters. The cases are certified to IP-67.


    Maximum protection and comfort system
    For maximum protection in cases Peli uses pre-cut expanded polyurethane foam (polyurethane foam).The cut foam is convenient for stacking and fitting to specific equipment sizes. We offer a choice of cases in various configurations:
    – foam plastic with a notch;
    – universal modular partitions, which are dividers inside the bottom of the case (with Velcro fasteners) and are located at your discretion.

    Numerous retrofit options
    PELI ™ cases can be used to mount almost any device in them, therefore manufacturers of various equipment use PELI ™ cases as a ready-made instrument case.Installation of equipment inside the cases is carried out using a special set of fasteners. For the precise design of the future device, you can freely download the drawings and CAD files of cases and mounting frames on the manufacturer’s website.


    Celestron ExploraScope User Manual – Manuals +

    USER MANUAL

    WHAT’S IN THE BOX

    We recommend keeping the telescope box so that it can be used to store the telescope when not in use.Unpack the box carefully as some of the parts are small. Use the parts list below to check that all parts and accessories are included.

    PARTS LIST

    INCLUDED BUT NOT PICTURED.

    1. Eyepiece 4 mm
    2. 3x Barlow lens
    3. DVD-ROM TheSkyX First Light Edition
    SOLAR ALARM

    Never look directly at the Sun with the naked eye or through a telescope unless you have a suitable solar filter. This can cause permanent and irreversible eye damage. Never use a telescope to project an image of the sun onto any surface. Internal heat build-up can damage the telescope and any accessories attached to it.
    Never use an eyepiece solar filter or Herschel wedge. Heat build-up inside the telescope can crack or break these devices, allowing unfiltered sunlight to pass to the eye.
    Do not leave the telescope unattended, especially with children or adults unfamiliar with the correct operating procedures for your telescope.

    TELESCOPE ASSEMBLY

    SLIDE

    The ExploraScope mount and tripod comes fully assembled for easy installation.

    1. Remove the tripod and mount from the box.
    2. Loosen the handles at the bottom of each leg and extend the inside of the leg to the desired length.Tighten the knobs to secure the legs, being careful not to over-tighten.
    3. Set the tripod upright and extend the tripod legs until the center bracket snaps into place.
    4. Place the threaded rod on the bottom of the accessory tray over the hole in the middle of the leg support bracket and rotate the entire tray clockwise until the tray is firmly against the leg support (Figure 1).
    5. Ensure that the leg pivot bolts are secure by tightening the black knurled nuts on the top of each leg.
    TELESCOPE TUBE

    To attach the telescope tube to the tripod and mount:

    1. Locate the delayed rod guide to the side of the U-mount. Loosen the set screw in the rail until the hole in the rail is blocked.
    2. While holding the telescopic tube over the mount, thread the slow action rod through the guide on the mount and carefully lower the hinges on the sides of the telescopic tube into the supports at the top of the U-post (fig.2).
    3. Tighten the set screw on the rail to hold the telescope in place.
    4. Insert the two telescope tube mounting bolts through the top of the U-mount and into the loops on the sides of the telescope tube. Do not overtighten (fig. 3).
      NOTE: The 70mm refractor is shown, but the Newtonian 114mm refractor attaches to the mount in the same way.
    STAR DIAGONAL

    (ONLY FOR 60 MM, 70 MM AND 80 MM REFRACTORS)

    The diagonal of the star attaches to the back of a refractive telescope, which contains a small mirror that reflects light at a 90 ° angle, providing a more comfortable view position.It also gives you a 100% correctly oriented image making it easy to use this telescope for daytime Earth observations.

    1. Remove the plugs from both sides of the diagonal.
    2. Loosen the set screws on the back of the focuser and remove the small dust cap.
    3. Insert the smaller tube diagonally into the back of the focuser and secure it by tightening the set screws (Figure 4).

    You can turn the diagonal to any position by loosening the set screws.

    EYES

    Your telescope comes with two eyepieces, a low power (20mm eyepiece) and a higher power (4mm eyepiece) eyepiece. Whenever you tune the telescope, always start with a 20mm eyepiece. After your target is centered in the 20mm eyepiece, you can switch to the higher magnification 4mm eyepiece for more detailed information. view.

    To attach the eyepiece to a 60 mm, 70 mm or 80 mm refractor:

    1. Loosen the set screws at the open end of the star diagonal.
    2. Insert the 20 mm silver eyepiece tube into the diagonal of the star (Fig. 5).
    3. Tighten the set screws to secure the eyepiece.
    4. To see the sharpest image possible, you need to focus by looking through the eyepiece and slowly turning the focus knobs until you find the sharpest image.

    To set the eyepiece to 114mm Newton:

    1. Loosen the two silver set screws on the focuser and remove the small plastic dust cap.
    2. Insert the silver tube at the bottom of the eyepiece into the focuser (fig. 6).
    3. Tighten the two set screws to secure the eyepiece.
    4. To see the sharpest image possible, you need to focus the eyepiece. To focus, look through the eyepiece and slowly rotate the focusing
      NOTE: The 114mm Newtonian comes with a dedicated 20mm eyepiece that gives you the correct orientation for daytime ground photography.viewing. 4mm is for astronomical observation and produces an inverted image. This is completely normal.
    STAR SEARCH
    The

    ExploraScope comes with a StarPointer red dot finder that you will use as an aiming tool when aiming the telescope at a target.
    To install StarPointer finder:

    1. Remove the two silver nuts from the threaded rods at the top of the tube next to the focuser.
    2. Place the two holes in the finder base above the threaded posts so that the finder base is flush with the telescope tube. Make sure the round finder window is pointing towards the end of the telescope that you will be aiming at.
    3. Screw the two nuts back onto the threaded posts (fig. 7).
    4. A small transparent plastic tab is inserted between the battery and the contact. This was done to prevent battery discharge during transport.Pull it out to make the battery work.
    STAR POINT SEARCH ALIGNMENT

    The StarPointer helps you aim the telescope by looking through the round glass window and covering the target with a red dot projected into the window.

    When first assembling the telescope, you need to align the StarPointer with the main telescope optics. While this step can be done at night, it is much easier during the day. Once you have finished aligning the finder, you do not have to repeat this step, unless the finder is bumped or dropped.

    To align StarPointer:

    1. Take the spyglass outside during the day. With the naked eye, look for an easily recognizable object, such as a street lamp, car license plate, or tall tree. The object should be as far away as possible, but not less than a quarter of a mile away.
    2. Remove the main dust cover from the telescope and make sure your 20mm eyepiece is installed.
    3. Move the telescope left and right or up and down until it points roughly to the object you selected in step 1.
    4. Look into the telescope through the 20mm eyepiece and manually move the telescope until the object of your choice is in the center of the view. If the image is blurry, gently rotate the focus knobs until it is in sharp focus.
      NOTE: The image in the telescope eyepiece will be upside down. This is perfectly normal for an astronomical telescope.
    5. Once the subject is in the center of your 20mm eyepiece, turn on the finder by turning the power switch knob fully clockwise.
    6. Position your head about 20 cm behind the finder, look through the round window and find a red dot. It will probably be close, but not from above, the object you see when you look through the XNUMX mm eyepiece.
    7. Without moving the telescope, use the two adjustment knobs on the side and below the StarPointer. One controls the movement of the point to the left and right, and the other controls the movement up and down. Adjust both until a red dot appears over the same object that you observe with a 20mm eyepiece.

    Now select a few other distant targets to practice aiming the telescope. Look through the StarPointer window and place a red dot on the target you are trying to view and make sure it is a 20mm scope eyepiece. When your StarPointer finder is aligned, your telescope is fully assembled and you are ready to observe!
    NOTE: Be sure to turn off the StarPointer when not in use to conserve battery power.

    YOUR FIRST SESSION OF ASTRONOMIC OBSERVATIONS

    MOON

    You are now ready to take out your telescope at night and make some real observations!

    Let’s start with the moon.It takes about a month to complete the Moon’s phase cycle, from New Moon to Full Moon and back. Try observing him at different points in this cycle.

    Although you can see the Moon anytime it is visible in the night sky, the best time to view is from two days after New Moon to several days before Full Moon. During this period, you will be able to see most of the craters and mountain ranges of the Moon. Refer to the calendar to find out when the next New Moon will be.

    1. With a clear view of the moon, mount the telescope with a 20 mm eyepiece.
    2. Turn on the StarPointer finder and look through it to find a red dot.
    3. Move the telescope until you can see the Moon through the finder window and the red dot is at the center of the Moon.
    4. Look through the 20 mm eyepiece. Turn the focus knobs gently to adjust the sharpness of the image.

    CONGRATULATIONS! YOU OBSERVED YOUR FIRST HEAVENLY OBJECT!

    To get closer to the view of the Moon, replace the 20 mm eyepiece with a 4 mm eyepiece.This will give you more magnification and the Moon will appear much larger. When changing eyepieces, you may need to adjust the focus knobs for the sharpest image possible.

    You can view many other celestial objects such as planets, star clusters and nebulae using the same basic technique.

    For more astronomical observations, visit
    ExploreTheSky.com

    There you will find comprehensive tips to help you get the most out of your new telescope, including:

    • How to Observe Planets
    • How to find and observe stars, binary stars, star clusters
    • How to Observe Deep Space Objects such as Nebulae and Galaxies
    • How to choose a good place for astronomical observations
    • How to assess the state of the sky
    CELESTRON
    LIMITED TWO YEAR WARRANTY

    A.The Celestron warrants your telescope to be free from defects in materials and workmanship for a period of two years. Celestron will repair or replace such product or part that, when checked by Celestron, reveals defects in material or workmanship. As a condition of Celestron’s commitment to repair or replace such a product, the product must be returned to Celestron with proof of purchase satisfactory to Celestron.

    B. A correct return authorization number must be obtained from Celestron prior to return.Call Celestron at (310) 328-9560 to receive a number that will appear on the outside of your shipping container.

    All returns must be accompanied by a written statement stating the owner’s name, address and daytime telephone number, and a brief description of any claimed defects. The replacement part or product becomes the property of Celestron.

    The customer is responsible for all shipping and insurance costs to and from the Celestron plant and must prepay such costs.
    Celestron will use reasonable efforts to repair or replace any telescope covered by this warranty within thirty days of receipt. If repair or replacement takes more than thirty days, Celestron will notify the customer. Celestron reserves the right to replace any discontinued product with a new product of comparable value and functionality.

    This warranty is void and void if the product covered by the warranty has been altered in design or function, or has undergone mishandling, misuse, mishandling or unauthorized repair.In addition, this warranty does not cover product malfunction or wear due to normal wear and tear.
    CELESTRON DISCLAIMS ANY WARRANTY, EXPRESS OR IMPLIED, REGARDING MERCHANTABILITY FOR A PARTICULAR PURPOSE EXCEPT AS STATED HEREIN. CELESTRON’S SOLE UNDER THIS LIMITED WARRANTY IS TO REPAIR OR REPLACE THE PURPOSED PRODUCT IN ACCORDANCE WITH THE TERMS SET FORTH HERE. CELESTRON EXPRESSLY DISCLAIMS ANY LOST PROFITS, GENERAL, SPECIAL, CONSEQUENTIAL OR CONSEQUENTIAL DAMAGES, WHICH COULD RESULT IN ANY BREACH OF ANY WARRANTY, ORDER ANY WARRANTY OR CONSEQUENTIAL DAMAGES.ANY IMPLIED WARRANTIES, WHICH CANNOT BE DISCLAIMED, ARE LIMITED TO TWO YEARS FROM THE DATE OF THE ORIGINAL RETAIL PURCHASE.

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    This warranty gives you specific legal rights and you may also have other rights that vary from state to state.
    Celestron reserves the right to change or discontinue any telescope model or style without prior notice.
    If you have a warranty problem or need help using your telescope, please contact:

    Celestron
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    2835 Columbia Street
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    Tel: (310) 328-9560
    Fax. (310) 212-5835
    Monday to Friday 8: 4 AM to 4:00 PM PST

    NOTE:
    This warranty is valid for US and Canadian customers who purchased this product from an authorized Celestron dealer in the United States or Canada.The warranty outside the United States and Canada is only valid for customers who purchased the product from a Celestron International Distributor or Authorized Celestron Dealer in a specific country. Please contact them for any warranty service.


    FCC NOTE This equipment has been tested and found to comply with the limits for a Class B digital device, pursuant to Part 15 of the FCC Rules. These limits are designed to provide reasonable protection against harmful interference in a residential installation.This equipment generates, uses and can radiate radio frequency energy and, if not installed and used in accordance with the instructions, may cause harmful interference to radio communications. However, there is no guarantee that interference will not occur in a particular installation. If this equipment does cause harmful interference to radio or television reception, which can be determined by turning the equipment off and on, the user is encouraged to try to correct the interference by one or more of the following measures:

    • Reorient or relocate the receiving antenna.
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      Product design and specifications are subject to change without notice. This product is designed and intended for use by people 14 years of age or older.


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    90,000 Chapter 6 BRAKES Their power is needed for more than just stopping. Sports driving technique

    Chapter 6

    BRAKES

    Their power is needed not only to stop

    If you have decided to master only one technique of controlling the motorcycle, select “braking”. Using the brakes correctly makes every ride not only more exciting but also safer.Again, the focus is on your right hand. The index and middle fingers – not without the help of the right foot, of course – decide your fate. They are the main ones, only the brain is more important than them.

    Learning to twist the throttle is easy, it is much more difficult to learn how to brake. You will be confirmed by thousands of riders who, having properly accelerated on a straight line, were not able to extinguish the speed before the next turn. Correct use of the brakes is the most important part of motorcycle handling.

    YOUR FRIEND, BRAKE

    No matter how many books you read, you will not learn how to slow down.You will have to choose the time and practice on your personal motorcycle. Yes, of course, you say to yourself day after day, you ought to somehow carve out an hour for this business. In the best case, one day you drive to an empty parking lot and brake several times. The trouble is, accidents don’t happen according to our plans. The sooner you learn how to brake, the sooner you will be prepared for emergencies. Perhaps the trouble will happen tomorrow. Are you ready for it?

    I have seen many drivers unable to stop. No, they overclocked perfectly, but as soon as it came to a halt, their results were extremely unstable. And it’s not about the type of bike. Much more important are the fingers and the leg, and also the brain that controls them. Again, smoothness is the key concept. Start slowing down gradually, feel the weight moving forward, and only then push down on the lever and pedal. Everything should be done quickly and in a controlled manner. Squeeze the lever, do not jerk it.

    I have witnessed many falls and can say that most of them occur in the first two months of riding a new motorcycle.Every motorcycle requires a different approach, and you must find that approach, otherwise you will be in big trouble.

    SKILL INSPECTION

    All the tips, tricks and techniques you find in this chapter are based on a myriad of training and braking tests I’ve done. I, like you, have always understood the importance of this skill, but it was only the first test I took that made me move from words to deeds. We then invited a group of motorcyclists to an empty parking lot, hoping to write an article for Motorcyclist later.It was there that I realized that stopping a motorcycle is much more difficult than stopping a car. And the main condition for success is not the bike model, tire brand or brake pad material. The most important thing is the skill of the driver. It is the possession of the skill that distinguishes the one who is able to stop stably with a minimum braking distance in any weather, from the one who closes his eyes and presses on the brakes. I have seen riders who were afraid of the brakes, or did not take their power into account, or relied too much on them. They all knew how to accelerate perfectly, and then did not know what to do with the achieved speed.

    So, one fine morning, we gathered eight motorcyclists of varying skill levels in a huge empty parking lot in Torrance, California. We asked them to get their bikes up to 40 mph and start braking as soon as they passed the orange cone. The morning began on a tragic note – the very first rider, level with the cone, grabbed the brake lever, like a child for ice cream. The front wheel blocked and slipped sideways, the motorcycle fell, the driver broke his collarbone. This incident served as a perfect illustration of the rule: squeeze the lever, do not jerk it.

    Then I saw how one of the subjects became so obsessed with the brakes that he forgot to release the gas. And the other constantly blocked the rear wheel and released both brakes in fright, increasing the braking distance. The sun rose higher and higher, and the seven remaining motorcyclists accelerated and braked over and over. By noon, everyone had improved their results – the braking distance was shortened, the braking became stable and controlled. And this added confidence to our drivers.

    All eight test subjects honestly studied at the motorcycle school and passed the examination for the right to drive a motorcycle, but only one of them regularly trained his skills. They all left the parking lot that morning, knowing the strength of their training. And they are not alone. I, too, having seen enough of their successes and seeing what the inability to slow down leads to, left there with one desire – to train. And as much as possible.

    WHAT TO TRAIN?

    Any professional shooter will tell you that you don’t need to pull the trigger, you need to push it smoothly. The same applies to the front brake lever. We learned from the last chapter that you only need to open the throttle a little at first in order to load the rear wheel and increase the contact area of ​​the rear tire.You need to brake the same way – at first, lightly, to move the weight forward, squeeze the front fork and load the front tire. After that, you can slow down more decisively. Without initial braking, the weight movement will be too sharp. And if you squeeze the brakes too harshly and aggressively, the front wheel will slip into a skid before the weight can move forward and load it. Front wheel skid almost always results in a fall. So, remember: first we move the weight forward, then we slow down.

    Take the time to practice progressive braking.Train it on every ride, every slowdown and stop. The asphalt is not always rough and the road is not always straight. Progressive braking will help you slow down on a wet road, or in the middle of a bend, and even in the middle of a wet bend. If you just hit the brake hard, you will most likely fall.

    ON THE EDGE OF THE SOUTH

    After initial braking, press the brake lever for real and feel how well your motorcycle stops. Listen to the front tire – at maximum deceleration it starts to squeal slightly.Concentrate on the effort of the fingers of your right hand. Over time, you can either briefly block the front wheel if you are riding a motorcycle with a long wheelbase and long reach, or lift the rear wheel off the ground if you have a short sport bike underneath. Release the front brake as soon as the wheel is locked. Practice letting go not completely, but just enough to unlock the wheel. If you release the lever completely out of fright, your stopping distance will increase.

    This technique is called edge braking.The edge beyond which the wheel locks is the limit of the bike’s braking properties in straight-line motion. You must master this technique perfectly.

    When braking with the rear brake, press down on the pedal with your toes, carefully controlling the force. Remember that the rear brake can be used, but very carefully. A locked rear wheel, especially in a corner, will at least scare you, and in the worst case, cause you to fall. Keep your foot on the footboard to avoid losing pedal feeling, just roll it over for more control over the effort.We’ll take a closer look at the rear brake application below.

    FRONT TIRE AND 100 CLUTCH UNITS

    Remember 100 clutch units? When driving in a straight line, the front tire can provide tremendous braking forces. That is why you can easily lift a sports bike in a stoppie.

    But as soon as the bike tilts in a turn, some of the units will have to be spent on resistance to centrifugal force. At the beginning of the turn, there will be few such units, but as the angle of inclination increases, their number will increase.Therefore, you will have to gradually release the brake lever to release the braking units. This technique is called trail-braking. If you do not release the lever, the number of requested units will become too large and the tire will slip.

    What exactly will happen next – demolition or fall – depends on the method of crossing the line. If the rider gains 100 units gradually, with progressive controlled efforts, then the result is a completely controlled drift. With sharp braking, a fall is inevitable.

    Anyone who has tried to turn under hard braking has noticed that this is not easy at all. If you release the brakes a little, letting the fork straighten a little, then it will be easier to turn. Think about this when you divide the units into “corner” and “brake”.

    It is not necessary to completely release the brakes when cornering. First, the speed may still be too fast for this turn. Secondly, the front fork will straighten out completely, reducing the bike’s steering, which is not necessary at all when cornering.The rider should gradually release the pressure on the lever as the incline increases in order to provide the front tire with a sufficient number of corner units. This is where the “sense of the lever” comes in handy, allowing you to accurately dose the effort and not go over the limit. The rider swaps out “brake” units that compress the fork for “angle” units that also compress the fork, and if done correctly, the motorcycle rewards you with smoothness and stability.

    TRAIL-BRAKING NEEDS GENTLE HANDS

    The motorcycle can turn and decelerate at the same time.This is called trail-braking. One day this technique will come in handy, for example, when in the middle of a turn you find yourself in an emergency. If in a corner you use 84 units to combat centrifugal force, you have 16 units to brake. And if you know how to use the front brake and load the tire smoothly, you can take advantage of those sixteen units without going over 100 units.

    As the bike decelerates, the lean angle for a given cornering radius will decrease, giving you more braking points.Use a simple equation: radius = speed. Decreasing the speed decreases the radius, or allows you to stay on track by decreasing the lean angle. But to put this equation into practice, you need to learn how to brake softly and in a controlled manner. Trail-braking is the answer to the question “what if I entered a corner too quickly?” Look where you want to go and slow down. As the speed decreases, the bike will straighten out, allowing you to brake even more. If the speed is too high – don’t give up, keep on braking.

    BRAKES AND OVERGASING

    In the second chapter we examined the basic techniques of controlling a motorcycle, with their help it is quite possible to move. But there are more advanced techniques that make our sport so exciting.

    Many people know how to brake, almost everyone knows how to change gears down, but to combine these two skills you will have to practice. In the end, why should you sacrifice braking for the sake of downshifting, or, on the contrary, wait with shifting until the end of braking? And again the right hand comes to our aid.

    The purpose of downshifting is to increase engine speed. This could be due to an approaching turn, or a hill, or overtaking. If you brake before a turn, you will have to shift down at the same time. In an advanced driving school, you will definitely be taught a technique called heel-toe, when you press your toes on the brake and simultaneously with your heel or the edge of your foot on the gas. The goal is to increase the revs to match the low gear. This technique makes downshifting smoother and conserves traction.

    If you are using only two fingers for braking, you can wrap the remaining three fingers around the throttle grip and twist it while shifting downward. The hardest part is to keep slowing down at the same time. At first, you will certainly be letting go and then grabbing the lever again, which of course will not like the front fork. And nevertheless, rebasing must be done, because when shifting down, the rear wheel can be blocked, and this is not necessary at all for you in a turn, even with a slight tilt.Whenever you hear a rear tire squeal after a downshift, be aware that the revs are out of sync with the new gear. The rear wheel has to spin the engine through the chain, gearbox, clutch and crankshaft. Isn’t it easier to throttle with the clutch depressed? This will benefit both you and the engine and transmission.

    Most modern motorcycles allow you to adjust the distance from the brake lever to the handlebars. Take this opportunity and adjust the position of the lever to the size of your hand.I noticed that for some reason most drivers push the lever too far, thus making it difficult for themselves the delicate work of the brakes.

    Downshifting with simultaneous braking is quite easy to learn from the point of view of coordination of movements. Difficulties begin when it comes to the time it takes to complete the technique. You need to gas when the clutch separates the engine and transmission. It is not necessary to squeeze the clutch lever all the way, use the ring finger as a stop. Don’t drop the clutch, release the lever smoothly and in a controlled manner – it can save your life if you make a wrong rpm or accidentally downshift two gears.

    REAR BRAKE

    Oh, yes, there is also a rear brake. The obvious problem with using it is shifting weight, right? When braking, the weight shifts forward, loading the front wheel and relieving the rear. And the unloaded wheel is easy to block with even a little effort. A modern sports bike, under heavy braking, tears off the rear wheel, so that it can be locked with almost no effort. It becomes clear why many riders do not use the rear brake at all.

    Research shows, however, that two brakes can stop faster than one. Many riders have found that pressing the rear brake a fraction of a second earlier than the front brake helps move the weight forward more smoothly. Aside from sportbikes and look at motorcycles with a longer wheelbase, which also often carry passengers, then using the rear brake does not seem entirely useless. In general, it doesn’t hurt to train your right leg either.

    TRY YOURSELF

    Set the rear suspension to the softest option. Ask your friend to hold the bike behind the wheel, and swing your rear a few times, trying to squeeze the suspension as low as possible.Notice how quickly it returns to neutral. Now ask a friend to apply the rear brake while you are compressing the suspension as much as possible. Have you noticed how much slower she is now straightening? This is because a clamped brake creates additional torque on the swingarm and frame. Thus, when using the rear brake, the front fork is compressed and the rear suspension is slower. I know many riders who use the rear brake just for this effect, because it allows better control of both braking and forward weight transfer.

    USE CAUTION ONLY

    Never treat the brake pedal of a motorcycle the same way you treat the brake pedal of a car. Usually, when the driver thinks that he is approaching a turn too quickly, he presses on the pedal with all his might, downright trample it. The result is well known – a black trail leading to the side of the road, and stories like “no, you need to take this turn more slowly”. A light touch on the pedal is a good way to dampen the speed before a corner, and a great way to adjust the speed directly into a corner.Why? Because applying the rear brake does not load the nose as much as applying the front brake. Of course, the nose will boot, but it will be much safer.

    In addition, the rear brake helps steering. Personally, I think this is its most important function.

    I have raced and tested motorcycles for many years and have come across machines without rear brakes on several occasions. It was once a tuning bike with an edgy carbon rim that looked smart but was completely dysfunctional. Another time, the pit stop mechanics in an endurance race were in such a hurry that they left me without rear pads. I also remember the unevenly screwed on the rear bracket holder, because of which the inner block was pressed against the disc and grinded off in several circles. And every time I felt that under heavy braking I was losing control of the bike, because as soon as the front fork clenched, he would start swinging his tail like a joyful dog. I felt like the bike was balancing on the front wheel, I didn’t feel the rear tire at all, so I was slow to turn every time.I’m not talking about trail-braking, the front brake cannot provide the same controlled and safe braking as the rear one. In short, I’m used to using the rear brake and it’s hard for me without it.

    I didn’t tell anyone about my love for the rear brake until I spoke to renowned racing mechanic Scotty Beach. We discussed new technologies with him, and I mentioned that I actively use the rear brake. Beach, on the other hand, spoke about Kurt Knoll, the famous ring stick. Kurt isn’t racing now, but you probably remember him if you saw him win all three Suzuki Cups in 1992. In addition, he has won numerous endurance races. In short, an experienced racer. Nor did he ever use the rear brake. One fine day, Beach began to describe the benefits of the rear brake with or without reason. After a while, Hall gave up and began to learn how to use it. As the skill grew, the lap times decreased, and since then Hall has applied the rear brake constantly.In general, no matter what you say, you can be sure that the fastest riders actively use the rear brake. Mick Duane, a five-time Grand Prix 500 champion, even asked for the rear brake actuator to be placed on the steering wheel when he was unable to pedal due to injury. Why would he need this if the rear brake is not very important?

    SLIP OR NOT?

    The rear brake is also an excellent device for measuring road grip. Push the pedal to the skid in a straight motion – the required effort will tell you how slippery it is now. I always check the canvas on rain trips this way. In addition, the properties of the new rubber can be assessed in this way. You just have to get used to the braking behavior of the rear tire, even the sound it makes. I repeat – such a test can only be carried out in a straight line with a reasonable speed and block the wheel only for a moment so that it does not slip sideways. And also – the softer you press the pedal, the more information you get.

    If you accidentally block the wheel, there are two ways to deal with it.First, you can release the brake pedal before the rear wheel moves to the side. The second way is taxiing. You leave the wheel locked and dampen the rear wheel vibrations. Steer so that the wheel is always straight to prevent the motorcycle from falling. Which is the best way? In an emergency, you are unlikely to think about the effort on the pedals, but at the same time, a trained right leg really shortens the stopping distance. So train both ways.

    ANTI-LOCK BRAKING AND COMBINATION SYSTEMS

    Anti-lock braking systems, or ABS, appeared on motorcycles in the 80s. They really reduce the likelihood of wheel locking when braking on difficult surfaces. The first time I encountered such a system was on the BMW K100 tests in Berlin. The organizers here and there covered the airport taxiways with sand and laid the route in such a way that braking began on the asphalt and continued on the sand. The first few inhibitions raised the pulse of journalists to the limit, but over time we realized that the system was working and trusted it.

    Since then, ABS systems have become more and more perfect.Is it worth buying such a system? Of course it is. Even on dry asphalt and hot tires, you need to be a master to beat electronics.

    Some motorcycles are equipped with combination systems where pressing the pedal or lever activates the pistons in both the front and rear braces. The combined system allows you to use the front brake and take advantage of the stabilizing effect provided by the rear brake. The bike is better at cornering and you can better control braking as your hand is more sensitive than your foot. In the event of emergency braking, you can use your leg as well, but in most cases, arm strength will suffice.

    REDUCTION OF EFFORT

    You will most likely want to adjust the rear brake for yourself. First, move the pedal down, away from the footrest so that your foot cannot easily reach it. This will help reduce pedaling effort. Just make sure that the pedal set in this way does not cling to the asphalt when cornering. And don’t forget to readjust the limit switch that turns on the brake light.

    If you continue to block the rear wheel after this, try reducing the efficiency of the disc-pad system. Some AMA Supersport Championship teams drill huge holes in the disc, some grind the abrasive layer off the pads. If you are not sure if you can handle it yourself, take the bike to a workshop. If the financial issue is not very acute, you can buy a special tuning bracket of a smaller size. This is the case when less is better. Either way, your job is to make the rear brake less responsive and therefore more useful. Remember, the purpose of the rear brake is not stopping, but stability and precise speed adjustments. If it blocks the wheel, it is useless.

    Just remember that no matter how well you own the rear brake, the front brake is the most important. Only he can save your life. The rear brake, on the other hand, enhances your driving experience, helps you to be less dependent on the weather and to achieve stability.

    RACING LESSONS

    Press down, then press

    The best riders try to slow down as much as possible before the corner at the very beginning of the braking zone.Once the front wheel is loaded, they push the lever with one powerful force. On the rest of the braking area, they fine-tune the speed with trail-braking, gradually releasing the braking force as the bike tilts.

    Accurately adjust your speed

    Riders use the brakes around the corner, all the way down to the apex, to get the most accurate cornering speed possible. Trail-braking helps the rider cope with incorrect corner entry speeds, or incorrect gears, or corner emergencies.The ability to control speed will enable stable passage through the apex, with or without traffic.

    Cleanliness is the key to health

    As I mentioned in the second chapter, mechanics often clean discs using mechanical or electrical tools. This should be done every week.

    Take your time with new pads

    New pads spoil the brake discs if used aggressively. Therefore, mechanics stick a sticker “new pads” on the tachometer so that the racers do not brake too much at first.

    Less is more

    Mechanics drill huge holes in the rear discs to make the bike lighter and reduce the effectiveness of the rear brake. Riders and regular drivers alike don’t need a powerful rear brake.

    Late braking? Forget!

    Aggressive braking on the streets is only appropriate in an emergency. Even in races, late braking is only used to change position, when the rider is ahead of the opponent before entering a turn and blocks his trajectory. This style degrades lap times because the bike has to ride within the ideal trajectory, which results in loss of apex, slower cornering speeds and imperfect straights.

    BUNCH 1 Trail-braking: Accurate speed selection

    Trail-braking increases safety because this technique allows you to change the speed in a corner. If you brake before a corner and release the brakes in a corner, then you have to decide too early how fast to take the corner.On a familiar route, this is not a problem, unless, of course, you come across some unpleasant surprise in the middle of the turn. But if you get caught, then you can’t do without the ability to brake in the slope. Remember, the faster you drive, the more you use the brakes, because you brake earlier and brake for longer.

    BOX 2 Basic Law of Training

    If you are accelerating to 120 mph, practice braking from 120 mph. You will find that stopping at this speed is very difficult.At the same time, pay attention to the length of the braking distance, perhaps this will force you to reconsider your speed limit.

    90,000 Weapons by Robert Hillberg. Part three
    Dear Readers! This is the third article in a series of publications devoted to weapons designed by American designer Robert Hillberg.

    In the previous installments, I introduced you to the Winchester Liberator and Colt Defender multi-barreled shotguns.

    Today I will introduce you to the COP .357 Derringer pistol.

    As strange as it may sound, the COP .357 Derringer is a direct descendant of the Winchester Liberator and Colt Defender shotguns. It became a logical development of the concept of multi-barreled weapons, proposed by Robert Hillberg in the mid-60s of the last century. This small pistol used the same technical solutions that were used and honed on Hillberg’s shotguns – with the only difference that our today’s guest has all the components in reduced sizes.

    It was rumored that, according to statistics, every 5th police officer in the United States is killed by his own weapon, which falls into the hands of criminals. Perhaps this is why, in the early 80s, Robert Hillberg pondered what should be a melee auxiliary weapon for police officers.

    And he reasoned, probably, something like this: the main properties that any weapon should have is its extreme simplicity and high reliability. (And on simplicity and reliability, he ate the dog, designing and modernizing all kinds of “Liberators” for all kinds of “Cheburator”.) In addition, taking into account the police specifics, in addition to the reliability of the action, the backup weapon must be adapted for concealed carrying, its sudden withdrawal and instant use.
    This is often a “last chance” weapon designed to be used at close range, so ideally self-defense should look like this: suddenly grab the weapon, pull the trigger and fire ahead.

    The revolver is suitable for these purposes, but not everyone is satisfied with its large dimensions due to the drum mechanism, so it makes sense to offer the police a product that has the advantages of a revolver, but devoid of its disadvantage associated with dimensions.

    Traditionally, Derringer-class pistols were considered the last-chance weapon. So why not try combining the benefits of the Derringer with the proven shotgun system?

    Some time later, a rather compact pistol called the C.O.P. which stands for “Compact Off-Duty Police”.
    A campaign called COP Incorporated was registered (now defunct) specifically for its production in Torrance, California.

    Derringer was distributed by another local campaign: M&N Distributors of Torrance.Since the pistol was designed for the powerful .357 Magnum revolving cartridge, its full name sounded like this: COP .357 Derringer. By the way, its design allowed to shoot with .38 Special cartridges without making any changes.

    COP .357 is a type of multi-barreled non-automatic pistol.
    The pistol consists of 54 parts. Its body is completely made of stainless steel.

    It is a four-shot weapon with a pair of four barrels in one block, made in the form of a single piece.
    Each barrel had its own separately installed firing pin.

    Pistol COP .357 with broken barrel block. Arrows indicate strikers.

    The barrel block is connected to the frame by means of a hinge in its lower rear part.

    Hinge joint for connecting the barrel and frame in the pistol COP. 357

    The pistol is loaded and reloaded manually, one cartridge at a time and is done by breaking the barrel block downwards, according to the principle of a double-barreled hunting rifle.The cartridge cases are partially ejected from the chamber by means of an ejector. Further, the sleeves are manually removed by the shooter one by one.

    How can I not recall the commentary to my first article in this series (about the Liberator):

    Quote: GRAY

    So this is where the “Wasp’s” legs grow from.

    The barrel block latch is located on the upper part of the pistol frame and, in addition, acts as a rear sight.

    Pistol Sights COP .357

    To open the lock and break the barrel block, pull the rear sight with your thumb. The upper surfaces of the pillar are grooved like the edge of a coin to prevent the finger from slipping.


    Place your thumb on the rear of the barrel and pull towards you to open the barrel assembly of the COP .357 gun.

    But the four barrels and the way of locking the barrel block are not the only similarities with the Winchester Liberator shotgun designed by Robert Hillberg. The COP .357 Derringer pistol uses a trigger mechanism that closely resembles that of the first generation Liberators.This is a Double Action Only trigger with a hidden trigger. That is, each time the trigger is pressed, the mechanism is first cocked, and then released and, accordingly, shot.

    Since the trigger force on COP .357 was “not childish”, many users admitted that they had never succeeded in firing a series of four shots: their fingers and wrist were very sore, and he had a lot of tension, and the recoil from the powerful cartridge was considerable. The sequence of shots occurred due to the fact that each time the trigger was pressed, the percussion mechanism turned 90 degrees and hit the next striker.

    Since it is generally accepted that a weapon with a trigger, firing only self-cocking, does not need a safety catch, because a shot can only occur when the trigger is fully squeezed, there was no manual safety catch in this pistol.

    In October 1983, patent US4407085 A for the main components of the pistol was received and published.

    There was also an opposing patent: publication US 1348035 A dated July 27, 1920. It was issued to Oscar Mossberg. The patent contained a description and drawings of the main components for a compact multi-barreled automatic pistol.On the basis of these units, a Mossberg Brownie pistol was assembled chambered for the .22 Long Rifle.

    The Mossberg Brownie pistol cost only $ 5 and was produced for 13 consecutive years (1919-1932). About 20 thousand copies were produced.

    COP .357 Derringer was produced in small quantities, but it did not gain much popularity either among the police or in the civilian market. Despite the fact that this pistol was approximately twice as wide and significantly heavier than any automatic pistol in. 25 ACP (6.35×15 mm Browning), its relatively compact dimensions and powerful cartridge made this pistol a good candidate for a last-chance weapon.

    The thickness of COP .357 is decent, and considering that it weighs 800 grams, even without cartridges we get a “lethal” weapon in the form of brass knuckles.

    Still from the movie Crying Freeman (1995). During the final showdown with the yakuza, a guy named Koh (Byron Mann)
    appears with a COP pistol.357, which was fed into the palm by a mechanism hidden in the sleeve. And although it is a four-shot, the hero fired from it 8 or even 9 times.

    COP .357 Derringer pistol in branded holster.

    According to unconfirmed information, they tried to fix the situation by releasing a version of COP Derringer chambered for the low-impulse .22 Winchester Magnum Rimfire cartridge of the 5.6 mm family, but it apparently did not work either, and production was curtailed.

    In 1990, its production was resumed by the American Derringer campaign, but due to the lack of sufficient buyers, its production was soon discontinued.

    At the moment, COP .357 pistols are quite rare and collectible. Their average market value, depending on the state, ranges from 900-1000 US dollars. They say that “strongly yuzanye” can be found cheaper: 200-350 bucks.

    Ads for the sale of COP .357 pistols posted on the board ArmList.COM

    Before I move on to describing the replicas and its roles in the films, I enclose brief specifications of the COP pistol.357 Derringer:

    Airsoft
    Renowned Airsoft replica manufacturer Marushin offers COP .357 Derringer pistols in 2 versions and 2 colors:

    Marushin COP .357 short barrel available in Silver and Black.
    Price $ 89.

    Marushin COP .357 Long Barrel also available in Silver and Black. Price $ 94.

    Cinema

    The COP .357 Derringer pistol was used as a props in the filming of films and television series, in anime and even in a computer game.

    First I want to highlight the work of Stephen Dane: he was assistant art director David Snyder during the filming of Blade Runner (1982). Before the filming of the film, this very assistant art director sketched a few sketches of some “weapon of the future”, which he proposed to equip the main character of the picture, Deckard (Harrison Ford). Here are the sketches:

    Soon the pistol itself was made according to these sketches, but most of them did not like it very much: it lacked “zest”.

    The first prototype of the blade runner weapon.

    Then a second prototype “blaster” was proposed for Deckard. Here it is:

    The second prototype of the Blade Runner weapon.

    Agree that they represent nothing more than a slightly modified COP .357 Derringer pistol. The second prototype already looked like a movie weapon should look, but it was also rejected (they say, Ridley Scott himself) and expelled from Rick Deckard’s arsenal, but they armed him with a “cross between a bulldog and a rhinoceros”: “Los Angeles Police Department – 2019 blaster ”, Which was assembled from the components of the“ Bulldog ”revolver from Charter Arms and the Steyr-Mannlicher Model SL rifle.

    Still from the film Blade Runner.

    LAPD 2019 blaster

    LAPD 2019 blaster

    Instead of this, Leon Kowalski (Brion James) was armed with a completely normal COP .357 Derringer pistol so that he would shoot another “runner” at the very beginning of the film: Holden ( Morgan Poll).

    Still from Blade Runner.

    Japanese replica based on COP .357. She is credited to Stephan Dane, the creator of the police transport for Blade Runner: Spinner Dokuhon.

    Here is a list of films that used the COP .357 Derringer pistol:

    Blade Runner (1982). Starring Harrison Ford.

    The Blue Iguana (1988).

    Blood In, Blood Out (1993).

    Bad Boys (1995). Starring Martin Lawrence and Will Smith.

    Crying Freeman (1995). Starring Mark Dacascos.

    The Matrix Reloaded (2003).Featuring Monica Bellucci.

    War (2007). Starring Jet Li and Jason State.

    21 Jump Street / Macho and nerd (2012).

    Black Out / Memory Blackouts (2012).

    TV series using the COP .357 Derringer pistol:
    Battlestar Galactica / Battlestar Galactica (2004).
    Stargate SG-1 / Stargate SG-1 (1997-2007).
    Special Unit 2 / Hunters for evil spirits (2001-2002).
    Psych / Clairvoyant (2006-2014).

    Anime using COP .357 Derringer pistol:
    Monster (2004-2005).

    A game that used a COP pistol .357 Derringer:
    Team Fortress 2.

    To be continued …

    Sources of information:
    http://en.wikipedia.org
    http://ru.wikipedia.org
    http://www.armslist.com
    http://www.guns.com
    http://zbroya.info
    http://props.steinschneider.com
    http://www.kitsune.addr.com
    http: // www.onpointsupply.com
    http://spec-naz.org
    http://weaponland.ru
    http://www.shooting-ua.com
    http://forum.guns.ru
    http: // world. guns.ru

    Neuroprotection using gene therapy to stimulate the expression of vascular endothelial growth factor – gene therapy

    items

    Abstract

    Engineered zinc finger protein (ZFP) transcription factors induce expression of endogenous genes and can be delivered remotely using adenoviral vectors. One such factor, Ad-32Ep65-Flag (Ad-p65), directs and induces the expression of vascular endothelial growth factor (VEGF; also called VEGF-A) splicing variants in their normal biological stoichiometry. We show that Ad-p65 transfection of primary motor neurons results in the expression of the VEGF variant and a significant increase in axonal growth in these cells. Given the neuroprotective effect of VEGF and its ability to increase neurite outgrowth, we investigated the efficacy of Ad-p65 to enhance motor neuron regeneration in vivo using rats that had suffered recurrent laryngeal nerve (RLN) injury.Injection of Ad-p65 after crushing the RLN accelerated the recovery of vocal cord mobility and the percentage of nerve contacts in the thyroaritenoid muscle. In general, adenoviral delivery of the engineered transcription factor ZFP, which induces the expression of the VEGF-A splicing variant, enhances nerve regeneration. Gene therapy for transcription factor ZFP to increase the expression of complete complement of VEGF-A splicing variants is a promising treatment for nerve damage and neurodegeneration.

    Entry

    Injury to the recurrent laryngeal nerve (RLN) leads to paresis or paralysis of the vocal fold. However, available treatments for vocal cord paralysis do not restore vocal cord movement, but rather provide geometric solutions, either by moving the paralyzed vocal cord closer to the midline to improve contact by avoiding obstacles, or using reinnervation to restore or maintain laryngeal muscle tone. 1, 2 The future ability to restore vocal cord movement for this dynamic neurological problem depends on strategies to maintain the survival of axotomized neurons and stimulate axonal regeneration in the corresponding laryngeal muscles.Treatment with neurotrophic growth factors such as vascular endothelial growth factor (VEGF; also called VEGF-A) is one approach that has the potential to preserve and restore functional innervation in axotomized vocal folds. Although VEGF was first characterized as an angiogenic factor, it is also known to have neurotrophic effects on the nervous system. VEGF is able to increase axonal outgrowth, 3, 4, by blocking neuronal apoptosis 5, 6, 7 and promoting neurogenesis. 8, 9 In addition, VEGF treatment is effective in numerous in vitro and in vivo neurodegeneration models such as amyotrophic lateral sclerosis and X-linked spinbulbar muscle atrophy. 10, 11, 12, 13

    In the nervous system, alternative splicing results in at least five different VEGF isoforms. 14, 15 Previous studies of the in vitro and in vivo efficacy of VEGF against neurodegeneration have focused on the efficacy of one VEGF variant, VEGF 165 . 10, 11, 12, 13 During angiogenesis, individual variants of VEGF splicing show different roles, and expression of multiple isoforms leads to a more robust angiogenic response. 16, 17, 18, 19, 20, 21 Therefore, the use of the full biological set of VEGF splicing variants may offer increased neuroprotection against nerve damage and neurodegeneration. 14, 16, 21 Engineered zinc finger protein (ZFP) transcription factors induce the expression of endogenous genes.Keyes and colleagues at Sangamo BioSciences Inc. (Richmond, CA, USA) developed a ZFP specific for VEGF 20, 22 under named 32Ep65-Flag, which induces the expression of VEGF and leads to the production of all naturally occurring splicing variants in their normal stoichiometry., 23 Expression in vitro and in vivo causes the activation of the VEGF variant and stimulates angiogenesis, improves blood flow and accelerates wound healing in animal models of diseases. 20, 22, 23, 24, 25, 26

    Due to problems with the introduction of growth factors into neurons 2, 27, 28, 29 delivery of viral vectors carrying transgenes encoding or inducing growth factors into damaged neurons is a viable alternative.Viral vectors can be delivered to the central nervous system (CNS) by remote injection into peripheral nerves, providing easier surgical accessibility, cell selectivity, and less risk of CNS damage than direct injection. 2, 27, 28 Our group and other researchers have established that these adenoviral vectors are transferred into the CNS by retrograde transport and lead to the expression of genes encoded by the virus. 27, 28, 30, 31 Delivery of neurotrophic growth factors such as insulin-like growth factor-I and gliotic neurotrophic factor was investigated for their ability to enhance regeneration in in vivo models of RLN injury. 30, 32, 33, 34 The efficacy of VEGF neuroprotection against RLN damage, however, has not yet been investigated.

    Considering the neuroprotective effects of VEGF treatment in vivo on neurodegeneration and the effectiveness of the induction of the full biological range of VEGF variants in ischemic animal models, we tried to determine whether the induction of VEGF expression provides neuroprotection in rodents that have undergone nerve compression injury in pn. This was accomplished using a ZFP transcription factor delivery adenoviral construct to enhance expression of VEGF splicing variants named Ad-32Ep65-Flag (Ad-p65). We show p65-Flag expression after transfection of primary motor neurons (MNs) and subsequent VEGF activation in these cells. Injection of Ad-p65 to rats with a nerve compression injury induces an earlier return of vocal cord mobility, suggesting increased nerve regeneration. This approach demonstrates the effectiveness of ZFPs for enhancing the expression of endogenous neuroprotective genes and holds promise for the treatment of vocal cord paralysis and other forms of nerve damage and neurodegeneration.

    Results

    Transfection of primary motor neurons and effects of increasing vascular factor endothelial regulation

    Primary MNs were transfected with Ad-p65 (250 μg (viral genomes) per MN) 1 day later in culture and fixed 48 hours after transfection for immunocytochemistry (ICC). MNs labeled with TUJ1 show successful transfection, as evidenced by the detection of the Flag epitope (Figures 1a and b). Successful transfection was further confirmed by the detection of the Flag epitope using Western blotting of cell lysates from the transfected MNs (Figure 1c). The up-regulation of VEGF variants after transfection with Ad-p65 was confirmed by Western blotting (FIG. 1d). Dot blot analysis of conditioned medium and enzyme immunoassay of cell lysates from transfected MNs also confirmed upregulation and secretion of VEGF from MN, with an approximately 86-fold increase in VEGF levels after transfection from 250 Vg to MN (Figure 1e; data not shown). To confirm that VEGF activation by transfection with Ad-p65 is capable of eliciting functional effects in MNs, we then investigated the effect of VEGF activation by Ad-p65 on axonal growth in MNs.To ensure that any increase in axon length was not influenced by cellular mechanisms, MNs were plated at a lower seeding density to allow for the growth of isolated MNs. Transfection with Ad-p65 resulted in a significant increase in axon length, with VEGF activation leading to a 21% increase in mean length compared to untransfected controls (Figure 2). Overall, these data show the successful activation of bioactive VEGF after transfection with Ad-p65 MNs.

    Ad-32Ep65-Flag (Ad-p65) transfection of primary motor neurons (MNs) and induction of vascular endothelial growth factor (VEGF).MN was transfected with Ad-p65 (250 μg per MN) for 24 hours and then cultured for an additional 48 hours before fixation. ICC shows successful transfection as evidenced by the expression of the Flag ( a and b ; red) in Tuj1-labeled MNs ( b ; green). ( c ) Western blot analysis of protein lysates showed expression of the Flag epitope in transfected MNs. ( d ) Western blot analysis of protein lysates shows the expression of a complete set of VEGF variants (arrows).( e ) Conditioned media from MNs transfected with Ad-p65 (250 μg per MN) and untransfected control MNs were analyzed using dot blotting to show VEGF activation and secretion. Recombinant VEGF (25 ng) was included as a positive control. A full-color version of this figure is available in the online gene therapy journal.

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    Effect of Ad-32Ep65-Flag (Ad-p65) transfection on axonal outgrowth in primary motor neurons (MNs).MN was transfected with Ad-p65 (250 μg per MN) for 24 hours and then cultured for an additional 48 hours prior to fixation for Tuj1 ICC. Images were collected to quantify axon length using GraphPad Prism software (GraphPad Software, Inc., La Jolla, CA, USA) from at least 30 individual MNs over three experiments per condition. 72 h after transfection with Ad-p65, there was a significant increase in the axon process by 21%, * P <0.05.The error bars represent sem

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    Recovery of vocal fold movement is accelerated with the injection of Ad-32Ep65-Flag.

    To determine the neuroprotective effect of Ad-p65 on MNs in vivo, we then examined the effect of Ad-p65 injection in rats that had suffered a right RLN crush injury. Crushing injury to the right RLN results in unilateral loss of movement of the vocal folds and loss of motor end plates in the thyroid-istenoid muscle. 39 Compression of RLN damage follows a reproducible time frame for recovery and does not result in the loss of MNs in the ambiguous core. 39 Direct laryngoscopy was performed on each animal before discontinuation after 3, 7, or 14 days to examine the correct movement of the vocal folds. Normal vocal cord movement was present prior to the initial surgical procedure, and all animals had direct paralysis of the right vocal fold in the immediate postoperative period based on observational assessment on direct laryngoscopy.Observation scores at 3, 7, and 14 days show an accelerated return of vocal cord movement (Table 1). In both the experimental and control groups, there was no return of movement of the vocal fold after 3 days. After 7 days, rats injected with Ad-p65 compared to control Ad-GFP showed a significant increase in vocal cord movement. Figure 3 shows the accelerated return of vocal fold movement in rats injected with Ad-p65 7 days after direct laryngoscopy. After 14 days, the function returned to normal in both groups. These data suggest that Ad-p65 restores vocal cord movement earlier than untreated rats injected with a control vector.

    Full size table

    Ad-32Ep65-Flag (Ad-p65) enhances recurrent laryngeal nerve (RLN) function. Sequential series of photographs from direct laryngoscopy 7 days after RLN injury in rats receiving control (Ad-GFP) or Ad-p65 injections.In normal rats, both the right (R) and left (L) vocal folds move towards or away from the midline. After injection of Ad-GFP, the right vocal cord remains paralyzed without movement for 7 days after RLN injury (upper series). After injection of Ad-p65, there is a return to the right vocal function with normal movement 7 days after crushing the RLN (lower series). A full-color version of this figure is available in the online gene therapy journal.

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    Ad-32Ep65-Flag Injection Accelerates Neural Plate Contact Recovery

    Rats sacrificed 3, 7 and 14 days after crushing and injection were then analyzed for the percentage of nerve-to-endplate contact (PNEC) for each group (FIG. 4). Similar to the results of direct laryngoscopy, there was no difference in PNEC between animals injected with Ad-p65 (28.6 ± 2.2%, N = 5) compared to a control adenoviral vector (26.1 ± 1.2%, N = 5) after 3 days. However, 7 days after crushing, rats receiving the Ad-p65 construct significantly increased PNEC (71.7 ± 1.6%, N = 5) compared to control rats (29.4 ± 1.6%; N = 4) … 14 days after the procedure, there were no significant differences in contact with the neural plate between the experimental (69, 3 ± 2, 1%, N = 4) and control (64, 2 ± 4, 3%, N = 4) groups, although there was a trend towards increased recovery.is present in Ad-p65 treated rats. These data show enhanced restoration of neural plate contact in rats that underwent RLN nerve injury after injection of Ad-p65.

    Ad-32Ep65-Flag (Ad-p65) accelerates the return of the neural plate contact. ( and ) Comparison of the percentage of neural plate contact (PNEC) for control and experimental rats 3, 7 and 14 days after injection. A significant difference between control (Ad-GFP) and experimental (Ad-p65) rats was observed at the 7-day time point, * P <0.001.The error bars are sem ( b and c ). Representative image of neural plate contact after crush injury after 7 days in rats receiving a control injection of the vector Ad-GFP ( b ) or Ad-p65 ( c ). Laryngeal sections were processed for acetylcholine staining (motor end plates, arrows) followed by neurofilament immunoperoxidase (NF) (nerve fibers, arrowheads). A full-color version of this figure is available in the online gene therapy journal.

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    discussion

    Engineering ZFP transcription factors have tremendous potential in the treatment of disease and injury due to their ability to regulate different cellular activities in combination with different functional domains. ZFPs containing the p65 domain of the transcription factor NF-κβ can be used to activate the transcription of target endogenous genes. Using this approach, Keyes and colleagues at Sangamo BioSciences have developed a dedicated VEGF ZFP named 32Ep65. 22 These studies analyzed the ability of this construct to increase the expression of bioactive VEGF and provide neuroprotection in an in vivo model of nerve injury. We show the expression of engineered transcription factor, activation of VEGF variants, and subsequent effects on axonal growth in transfected primary MNs. In addition, we show a significant acceleration in the recovery of vocal cord movement and return of neural plate contact with 32Ep65 treatment in rats that underwent experimental RLN crush injury.

    Endothelial vascular growth factor is an angiogenic protein that also has neurotrophic properties. VEGF can increase axonal outgrowth, block neuronal apoptosis, and promote neurogenesis. 3, 4, 5, 6, 7, 8, 9 In the nervous system, at least five different VEGF isoforms are generated from alternative splicing of a single gene, which includes VEGF 121 , VEGF 145 , VEGF 165 , VEGF 189 and VEGF 206 14, 15 VEGF 165 is the most commonly studied variant and has neuroprotective properties in various models of peripheral nerve disorders such as traumatic injury, diabetic neuropathy, sciatic nerve injury, spinal cord dissection and ischemic neuropathy. 26, 40, 41, 42, 43, 44 The efficacy of VEGF 165 for neurodegenerative disease has also been shown in in vitro and in vivo models of amyotrophic lateral sclerosis and a murine model of spinbulbar muscular atrophy. 10, 11, 12, 13 The complete biological array of VEGF isoforms, however, may offer increased potency. 14, 16, 21

    The roles of different variants of VEGF splicing have been previously studied for different effects on angiogenesis.Expression of the three major isoforms of VEGF, VEGF 121 , VEGF 165 and VEGF 189 , provides a more potent in vivo angiogenic response in a mouse model of ischemic hind limb compared to the expression of VEGF 165 alone. 21 In addition, similar studies comparing treatments with different ratios and combinations of different isoforms also imply that certain mixtures may provide improved efficacy and safety. 16 The vasculature resulting from this combination treatment not only exhibits accelerated angiogenesis, but also generates less permeable and more mature vasculature. 16, 20 These results are due to differential solubility, affinity for heparin, and the ability of multiple isoforms to activate various VEGF receptors, including VEGFR2. 45 Taken together, these results imply that expression of the full range of VEGF variants has the potential to improve neuroprotection.

    Expression of 32Ep65 ZFP promotes the production of all naturally occurring VEGF splice variants in their normal stoichiometry. 23 In vivo 32Ep65 induces VEGF expression, stimulates angiogenesis and accelerates wound healing. 20 Several additional studies support the use of ZFP induction of VEGF for the stimulation of angiogenesis as well as for the treatment of experimentally induced peripheral diabetic neuropathy. 24, 25, 26 These data support the idea that engineered ZFP transcription factors are able to regulate gene expression in vivo and have a therapeutic effect 20, 23, 46 and support the use of 32Ep65 for the treatment of RLN nerve shift damage in current research. ,

    We use the adenoviral construct Ad-p65 to deliver 32Ep65 labeled with the Flag epitope to the nervous system. Delivery of adenovirus for gene therapy bypasses many of the limitations of direct pharmacological therapy with neurotrophic factors in neurological diseases and nerve injuries. Most neurotrophic factors, such as VEGF, are significant, do not cross the blood-brain barrier, and have poor diffusion capacity when injected directly into the CNS. 2, 27, 28, 29 Adenoviral delivery also offers the possibility of remote injection into peripheral nerves or muscles, as viral vectors are transported into the CNS by retrograde transport, thereby providing easier surgical accessibility, cellular selectivity and less risk of CNS damage. 27, 28 We show the successful expression of 32Ep65 after MN transfection with Ad-p65 (250 μg per MN), as evidenced by the detection of the Flag epitope in MN by ICC (Figures 1a and c).This subsequently triggers the activation of VEGF variants (Figure 1d) that are released by MNs (Figure 1e) and increase axonal growth in transfected MNs (Figure 2). These data are consistent with the literature and further support the efficacy of 32Ep65 in inducing VEGF and enhancing axonal growth. 3, 4, 20

    Gene therapy with neurotrophic factors such as insulin-like growth factor-I and gliotic neurotrophic factor has been previously investigated and promotes recovery from experimental RLN injury. 30, 32, 33, 34 These studies are the first to show the efficacy of VEGF variant activation for RLN damage. The RLN crush injury model is a viable model for the study of therapeutic interventions and nerve injury. 27 The crushing injury of the RLN retains the neural structure needed to transport therapeutic agents along the entire axon, allowing for remote administration and retrograde transport. In addition, it is easy to replicate using standard surgical procedures, ultimately providing a predictable sequence of denervation followed by reinnervation, which is associated with sprouting of axons in the preserved neural structure. 30 Results from our laboratory show a reproducible time course for reinnervation and recovery from nervous system injury in rats who have established reinnervation beginning between weeks 1 and 2 after injury and reaching completion by week 3. 39 Reinnervation is measured by PNEC calculation and subsequent functional recovery is measured by direct laryngoscopy. The level of reinnervation achieved at 3 weeks is comparable to levels in control (non-squashed) nerves as previously reported at 86.8% PNEC. 39 Our data confirm this timeline for control rats in which we see a return of vocal cord movement and an increase in PNEC at 14 days (Table 1; Figure 4), which is reminiscent of previously reported PNEC levels. 39 Experimental rats, on the other hand, showed significant recovery at 7 days in both PNEC and vocal cord motility, indicating a significant increase in recovery rate in rats treated with Ad-p65 (Table 1; Figures 3 and 4).

    Injection of adenoviral constructs into the RLN after crush injury promotes the expression of virus-encoded genes in the MNs of the ambiguus nucleus. 27, 28, 30 Based on our in vitro results showing VEGF secretion from MNs (Figures 1e and f), it is possible that, in addition to the direct neuroprotective effects of VEGF on MNs, the beneficial effects of Ad-p65 treatment could be mediated through the effect of VEGF on cells near the injection site or adjacent cells in the MN microenvironment.VEGF stimulates survival and invasion of Schwann cells, which may play a role in axonal regeneration. 47, 48, 49 Schwann cells, MN axons and endoneurial cells express VEGF receptors, further suggesting the possibility that up-regulation of VEGF by Ad-p65 may affect several cell types and contribute to neuroprotection. 41, 47, 48, 49 Additional studies are required to determine the exact mechanisms and contributions of cell types affected by VEGF in protection against nerve damage and neurodegeneration following Ad-p65 gene therapy.

    Although nerve crush is an excellent experimental model, more research is needed 39, 50, 51 in other models of nerve damage and neurodegeneration to establish activation of VEGF variants using ZFP gene therapy as an effective model for treating neurodegeneration associated with both injuries . .. and illness. In addition, delivery of 32Ep65 adenovirus may be limited by the limited duration of expression of the constructs.For example, the loss of significance between the 7- and 14-day control and experimental groups could be the result of decreased expression of the ZFP transcription factor. Future studies using an adenovirus-associated virus that is able to overcome the host immune response and is reported to have limited toxic effects and prolonged gene expression in vivo, 52 could potentially overcome these limitations.

    Overall, these studies support the use of gene therapy using 32Ep65 ZFP to induce a full biological range of VEGF splicing variants and improve neuronal recovery after RLN crush injury.We show successful in vitro transfection of MNs following viral transfection with Ad-p65 and subsequent VEGF activation, which has a beneficial effect on axonal growth. In vivo treatment in rats with Ad-p65 after RLN crush injury accelerates vocal fold motility recovery and increases PNEC 7 days after crush injury. These data support further study of gene therapy using 32Ep65 ZFP to activate the full biological suite of VEGF variants for the treatment of nerve damage and neurodegeneration.

    materials and methods

    Primary culture of motor neurons

    Primary MNs were isolated from Esprague-Dawley E15 rat embryos according to our previously published protocol. 35 For the cultivation of primary MNs, the MN density was adjusted to 2 × 10 4 and 2 × 10 6 MN ml -1, and 50 μl of the suspension were applied to coverslips coated with poly-L-lysine in 24 well plate. or 500 μl of the suspension was applied to poly-L-lysine coated 12 well plates.After 24 h, MNs were fed with a nutrient medium.

    Viral transfection of primary motor neurons and analysis

    Ad-32Ep65-Flag was donated by Sangamo BioSciences Inc. Viral transfection of primary MNs was performed as previously described. Briefly, virus (0-250 Vg per MN) was added 24 hours later to culture and MN incubated for 48 hours before fixing for ICC and harvesting cell lysates to show successful MN transfection, or harvesting conditioned media to demonstrate VEGF expression. ,

    For ICC, cells were plated at 50 μl -2 onto glass coverslips coated with poly-D-lysine, followed by fixation in 4% PFA for 5 minutes. Immunostaining was performed according to standard protocols as previously described. 37 Primary antibodies used include Flag (Sigma-Aldrich Co., St. Louis, MO, USA) and TUJ1 (Neuromics, Edina, Minn., USA). Cells were then incubated in a suitable Alexaflor-conjugated secondary antibody (Molecular Probes, Invitrogen, Carlsbad, CA, USA) followed by installation in ProLong Gold (Molecular Probes, Invitrogen).Images were collected using an Olympus BX-51 microscope. Neurite length was quantified using ImageJ software (ImageJ, US National Institutes of Health, Bethesda, MD, USA, //rsb.info.nih.gov/ij/). Ten images per condition were taken randomly in three separate experiments, for a total of at least 30 MN per condition. The length of the axon was traced from the periphery of the cell body to the tip of the axon.

    Conditioned media from MNs and cell lysates were harvested 48 hours after transfection and Western blotting, dot blotting, or ELISA was performed as previously described. Briefly, for Western blotting, cell lysates were prepared by scraping cells in RIPA buffer (20 mM Tris, pH 7, 4), 150 mM NaCl, 1 mM EDTA, 0.1% SDS, 1 mM Na deoxycholate, 1% Triton. X-100, 0.1% trypsin units per μl of aprotinin, 10 mg / ml of leupeptin and 50 mg / ml of PMSF. Equal amounts of protein were loaded into each lane of 12.5% ​​polyacrylamide gel and separated by electrophoresis prior to transfer to nitrocellulose membranes. For dot blotting, protein was equilibrated and loaded onto nitrocellulose membranes.Dot blot and Western blot membranes were incubated with the primary antibody overnight at 4 ° C and incubated with the corresponding secondary antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA, USA) for 1 hour at room temperature … Primary antibodies included VEGF (Santa Cruz Biotechnology) and Flag (Sigma). Antibody binding was determined using enhanced chemiluminescence with LumiGLO plus peroxide reagent (Cell Signaling, Danvers, MA, USA). Results were obtained using the BioRad ChemiDoc XRS Imaging System with Number One analysis software (Hercules, CA, USA). An enzyme-linked immunosorbent assay was performed using the PEGROTech Human VEGF ELISA Development Kit according to the manufacturer’s instructions (PeproTech Inc., Rocky Hill, NJ, USA). Results are representative of at least three independent experiments.

    Animal surgery

    All animals used for these studies were kept in a pathogen-free environment and ensured that the guidelines of the University of Michigan Animal Care and Use Committee were followed.Animal care standards outlined in the National Institutes of Health Laboratory Animal Care and Use Guidelines are followed to provide limited discomfort and stress.

    Thirty Sprague-Dawley rats weighing 300-350 g were subjected to inhalation anesthesia with isoflurane (Baxter Caribe Inc., Deerfield, IL, USA) followed by retraction of the anterior tongue to reveal the epiglottis. After confirmation of bilateral symmetric movement of the vocal fold by direct laryngoscopy, intubation was performed using a 16-gauge angiocatheter. General anesthesia was maintained using a Harvard Rat Ventilator (Harvard Apparatus, Holliston, MA, USA) and supplemental oxygen. After sterile preparation of the skin on the front of the neck, a vertical incision was made in the cervix, separating the muscles of the platysma. The submandibular glands and belt muscles were identified and separated to reveal the larynx, trachea, and thyroid gland. Using a dissecting microscope (Stereozoom 6, Leica, Buffalo, NY, USA), the right RLN was exposed and incised circumferentially from the middle trachea to the inferior border of the right thyroid gland.Then the nerve was crushed with jewelry forceps at the level of the sixth tracheal ring for 30 s. Following the injection of the virus (described below), the belt muscles and submandibular glands were re-approximated, followed by skin closure with staples. Direct laryngoscopy was performed again at the end of the crush / injection procedure and just before killing to quantify the movement disorder of the right vocal fold. During suspension, movement of the vocal fold was observed and movement was scored on a scale from 0 to 1 (0 indicates no movement of the ipsilateral fold, 1 indicates normal movement with complete closure of the glottis).Direct laryngoscopy was performed in a blind manner by one of the researchers.

    The study began with five rats in each group for the initial procedure as above. In the 7-day group, one experimental rat died on the 3rd day after surgery, presumably from a wound infection. In the 14 day group, one rat in the experimental and control groups was lost during postoperative direct laryngoscopy.

    Viral injection

    Rats were randomly assigned to six groups – three control groups and three experimental groups.The different time points for killing were the only differences between groups in the control or experimental study group. The control adenoviral vector (Ad / CMV-GFP; Ad-GFP) was injected into the RLN at a concentration of 1.2 × 10 7 pfu to the control animals that suffered from crush injuries. Animals in the three experimental groups were injected with RLN after crush injury using Ad-p65 at 0.9 x 10 7 pfu. 31 At in particular, after the identification of the nerve, a 3-0 silk suture was placed around the nerve and a slight resistance was applied.A 50 µm conical micropipette (glass micropipette remover PP-83; Narishige, Tokyo, Japan) was advanced under the perineurium along the nerve proximal to the site of crush injury. Using a microinjector and an oocyte micromanipulator (Nanoject; Drummond, Broomall, PA, USA), 1.5 μL of virus was injected into 50 nL boluses (5% volume error). This dose leads to maximum expression of the adenoviral transgene. 38

    Tissue processing and analysis

    Rats were sacrificed 3, 7 or 14 days after crushing and injection.The injection of Fatal-Plus solution (Vortech Pharmaceuticals, Dearborn, Michigan, USA) was followed by intracardiac perfusion with buffered 4% paraformaldehyde. Laryngectomy was performed followed by cryoprotection in phosphate buffer with 20% sucrose and insertion into a Tissue-Tek OCT connection (Sakura Finetek USA Inc. , Torrance, CA, USA). Laryngectomy tissue samples were cryosected in 20 μm axial slices along the long axis of the thyroaritenoid muscle. Cryosections (20 μm) of thyroaritenoid muscle were stained with acetylcholinesterase to identify motor end plates, followed by immunoperoxidase of neurofilaments to identify nerve fibers, as previously described. 27 Images were acquired using an Olympus BX-51 microscope (Olympus America, Inc., Center Valley, PA, USA).

    Statistical analysis

    The statistical significance between the control and the length of the MN axon transfected with Ad-p65 was calculated using ANOVA followed by Tukey’s multiple comparison test and standard t-tests (GraphPad Software Inc., La Jolla, CA, USA). PNEC in thyroaritenoid muscle sections was determined as an objective measure of the state of innervation by counting the total number of motor end plates in contact with neurofilament divided by the total number of neural end plates in each section. The count was blinded by light microscopy by one of the investigators and verified by the second investigator, and the results were averaged. Approximately every seventh section was analyzed. Analysis of variance using the Bonferroni multiple comparison test and standard t-tests was used to compare the average percentage of neurofilament / neural plate contacts between rats in the same group. A P value <0.05 was considered significant.

    Conflicts of interest

    The authors declare no conflicts of interest whatsoever.

    .

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