When Suturing The Skin Of Rats And Mice, What Type Of Suture Pattern Is Recommended?

Curr Protoc Immunol. Author manuscript; available in PMC 2008 Nov 25.

Published in terminal edited course as:

PMCID: PMC2587003

NIHMSID: NIHMS78703

TECHNIQUES IN Aseptic RODENT SURGERY

Shelley Fifty. Hoogstraten-Miller

¹ National Human Genome Inquiry Found, Bethesda, Maryland

Patricia A. Brown

² Office of Laboratory Animal Welfare, Bethesda, Maryland

Supplementary Materials

gloving movie: Video 1.12.1 Donning sterile surgical gloves.

GUID: 8F9D2738-A0B1-4955-BC27-7D4591E88603

Abstract

Performing aseptic survival surgery in rodents can be challenging. This unit of measurement describes some basic principles to assistance clinicians, researchers, and technicians in condign proficient in performing aseptic rodent surgery.

Key Terms: aseptic surgery, technique, rodent, surgical training, musical instrument preparation, suture material, anesthesia, analgesia, surgical gloves

Unit Introduction

Performing hygienic survival surgery in rodents tin can exist challenging. Unlike in larger species where there is a dedicated surgical suite and several personnel to help the surgeon, rodent surgery is most commonly performed alone. This means the surgeon must induce, maintain, and recover the creature from anesthesia, as well as, surgically prepare the animal, and perform the surgery aseptically.

The following principles described in the Guide (NAS, 1996) apply to rodent surgery:

1. Appropriate pre-operative and mail service-operative care of animals in accordance with established veterinarian medical and nursing practices is required.

2. All survival surgery will be performed by using aseptic procedures, including sterile gloves, masks, sterile instruments, and aseptic techniques.

3. A dedicated surgical facility is non required for rodents but surgery must be performed using aseptic techniques.

4. Research personnel will be appropriately qualified and trained in all procedures to ensure that good surgical technique is skilful.

Good surgical technique includes asepsis, gentle tissue handling, minimal dissection of tissue, appropriate employ of instruments, effective hemostasis, and correct employ of suture materials and patterns.

Aseptic surgery is performed using procedures that limit microbial contamination so that significant infection or suppuration does not occur. This includes training of the animal, preparation of the surgeon, sterilization of instruments, supplies, and implanted materials, and the use of operative techniques to reduce the likelihood of infection (NIH ARAC, 2005).

Strategic Planning

Aseptic technique is achieved, in part, by the pre-surgical planning that begins during protocol development in consultation with your veterinarian. This includes identification of personnel, their roles and grooming needs, equipment and supplies required for the procedures planned, the location and nature of the facilities in which the procedures will be conducted, and pre- and post-operative care.

i. Choosing the Surgical Area

Characteristics of a good surgical expanse include an uncluttered surface area that is easily organized and disinfected, and gratis of debris and equipment not related to surgery (figure 1.12.1). The area should exist defended for the duration of the procedure, merely can exist used for other purposes when non being used for surgery. Avert locations that are beneath supply ducts to minimize contagion from dust. Avoid high traffic areas such as those nigh doorways to prevent unnecessary interruptions and creation of air turbulence and contamination of the surgical field.

Good surgical site (left) and poor surgical site (right).

2. Considerations for Anesthesia and Analgesia, an Introduction (See Unit i.4)

Select the anesthetic based on the type of surgical procedure, the length of the surgical procedure, the equipment available and the expertise of those who will be responsible for administering the coldhearted (Flecknell, 1996; Kohn, 1997; Swindle, 2002). Consideration must too exist given to the application of pre-, intra-, and post-operative analgesia. Analgesics can be injected, applied topically in a drop-wise fashion to the surgical expanse and/or supplied in the food or water.

Inhalant gas anesthetics are administered using precision calibrated vaporizers (Figure 1.12.2). When using gas anesthetics you must account for scavenging of waste gases. I acceptable method of scavenging is the use of a downdraft table (Figure 1.12.3). It is important not to completely cover the surface of the downdraft table. This will cause a loss in its power to effectively scavenge gases. Downdraft tables are ordinarily only constructive up to a height of 6–8 inches from the surface. Do non utilise induction chambers taller than this for induction of anesthesia. Placing the chamber in a chemical fume hood (Figure 1.12.iv) or a type IIB biosafety cabinet (Figure 1.12.five) that is vented to the exterior are other methods than can be used to scavenge waste anesthetic gases. A charcoal canister (Figure 1.12.vi) attached to the part of the breathing circuit for expired gases can likewise be used for scavenging. Charcoal canisters must be weighed earlier, and after, each use and must be replaced subsequently an increase in the recommended weight. Depending on the size of the canister and the manufacturer'southward recommendations, the canister should as well be weighed during especially long procedures to clinch its continued effectiveness.

Anesthesia machine with precision calibrated vaporizer.

Type IIB biosafety cabinet.

Injectable anesthetics are widely used in rodent surgeries (e.g., ketamine/xylazine mixtures, pentobarbital, tribromoethanol). Controlled substances (pentobarbital, ketamine) require additional record keeping. If using injectable anesthetics, it is important to weigh each animal and dose each according to its body weight.

Some anesthetics, such as ketamine, cancel the blink reflex. Anesthetized animals should have their corneas protected with an ophthalmic lubricant. To avoid contamination of the lubricant, practise not touch the tip of the tube to the skin or eye surface.

Anesthetized animals must be monitored during the process to assure that they stay in the proper coldhearted plane. Do not allow them to get too light or too deep. Once the anesthetic has been given time to take effect, the anesthetic plane can exist assessed by pinching the toes, tail or ear of the animate being. Whatsoever reaction from the animal indicates that the animal is also light and that additional coldhearted should be given.

Monitoring should include inspection of the mucous membranes and exposed tissues. This will give an indication of tissue perfusion and oxygenation. The color should be a bright pinkish to reddish and not dusky gray or blue. Respiratory pattern and frequency is likewise easily monitored and will requite an indication of anesthetic depth and other potential complications.

Various types of instrumentation tin can assist in monitoring the anesthetized patient. Core body temperature tin can be monitored in rodents, including mice. Pulse oximetry, capnography, and electrocardiograms can be used in larger rodents to monitor pulse, oxygenation, and centre rate. Monitoring instruments must be properly calibrated, as inaccurate information may be misleading and could result in a compromised condition or fatalities.

The almost frequent complexity of small-scale animal anesthesia is hypothermia resulting in prolonged recovery or death of the animal. Animals should be provided with a estrus source during the pre-operative, intra-operative, and postoperative periods. Because of the loftier airflow, the risk of hypothermia is heightened when using downdraft tables, chemical smoke hoods, or biosafety cabinets.

The safest devices for providing heat to anesthetized animals are circulating hot water blankets or instant heat devices. These devices must exist covered with a paper towel or other insulation so that the animal does not come up in straight contact with the hot surface. Slide warmers tin also be used equally a rut source during recovery. By placing the recovery cage on the slide warmer information technology will exist pre-warmed and set up to accept the animal once the surgery is consummate. Use a thermometer to measure the temperature at the level of the animal. The temperature non exceed 85 to 95°F (29.4 to 32.2°C). Rut lamps and electrical heating pads can be very unsafe and should be used with corking caution.

three. Instrument Preparation

Planning the surgical procedure requires consideration of the instruments required for the procedure and what method of instrument sterilization will be used. In that location are 3 commonly used methods for musical instrument sterilization:

1. Steam autoclave or ethylene oxide. When using one of these methods a simple paper peel pack (Figure 1.12.7) or a complex pack (Figure i.12.eight) is used. A simple pare pack contains pocket-size numbers of small to medium sized instruments. A complex pack consists of overlapping cloth or paper drapes folded together and sealed with indicator tape. It can incorporate a big collection of instruments of various sizes (Knecht, 1987). Tip protectors should be added to frail instruments or those with sharp points. Delicate instruments, materials for implantation such as catheters or items that otherwise may melt or get damaged when heated can exist sterilized using ethylene oxide. The packs must exist sufficiently aerated to prevent toxic effects from rest gas. This may require 24–72 hours.

   

   

2. Cold sterilization. Glutaraldehyde solutions are effective if instruments are exposed for the proper length of time and expiration dates of prepared solutions are observed (unremarkably 28 to 30 days).

3. Dry estrus sterilization. Hot bead sterilizers (Figure 1.12.9) sterilize merely the tips of the instruments. The beads must exist pre-heated to the recommended temperature and the instruments exposed for the recommended fourth dimension. "Flash" dry estrus sterilizers (Figure 1.12.10) sterilize the unabridged instrument and as well requires adherence to the recommended temperature and exposure time. For both methods, gross debris must be removed from the musical instrument before sterilizing and the instruments must be allowed to cool before touching tissues.

   

   

   Flash dry out rut sterilizer.

Booze provides disinfection not sterilization and should not be used to sterilize instruments.

four. Option of Wound Closure Materials

The selection of the type and size of suture material should be done in advance of the surgical procedure. A 3-0 suture thickness or smaller is best. Cutting and reverse cutting needles accept abrupt edges and are best used for skin suturing. Not-cutting, taper or round needles are used for suturing easily torn tissues such equally peritoneum, muscle or intestine.

If ligation of vessels or suturing of tissues other than skin is necessary during surgery, an absorbable material such every bit polyglactin 910, polyglycolic acid, polydioxanone, polyglyconate, or chromic gut should be used. For pare closure, non-absorbable suture such as polypropylene, nylon, stainless steel wound clips or staples may be used. Well-nigh rodents volition gnaw at any externalized sutures, so a buried suture line or wound clips are recommended. Cyanoacrylate surgical adhesives may exist used to close incisions or to shut the area between sutures. Silk is a non-absorbable suture cloth that tin can cause tissue reactions and may wick microorganisms into the wound. Information technology is best used for cardiovascular procedures merely and not for closure (Knecht, 1987).

Basic Protocol ane: Surgical Preparation of the Animal

Once everything for the surgery is pre-selected and organized, the brute tin be anesthetized and surgically prepped. Surgical preparation includes removal of the hair and disinfection of the surgical site. Surgical grooming of the animal should occur in a location unlike than that used for performing the surgeries. This will help to foreclose hair and dander from getting on the sterile packs. If space constraints or requirements for employ of the down draft table, chemical fume hood, or biosafety chiffonier necessitates a unmarried location for prepping and surgery, and so the bench towel used to prep the beast should be replaced before performing the surgery. The surgical pack, if already open, must be covered with a sterile drapery to prevent contagion with hair.

Materials

"Mini clipper" with no. 0000 blade

Gauze pads (two"×2")

Adhesive record (1–ii" wide)

Cotton-tipped applicators

70% alcohol

Iodophor or chlorhexidine scrub

1. Using the clipper remove the hair from the surgical site.

   In mice, an easy culling to clipping the fur is to remove it past plucking. Hair follicles in mice are usually in telogen or resting phase, and hair tin can be removed without injury. It is a fast and easy method that does not go out stubble.

2. Dab the clipped or plucked area with a slice of adhesive record or moistened gauze to pick upwardly loose pilus that could otherwise migrate into the incision.

3. Utilise a gauze pad or cotton fiber-tipped applicator to prep the surgical site with alternating scrubs of an iodophor and 70% alcohol. Employ a circular movement beginning at the center of the shaved area and working toward the periphery. . Never become back to the center with the same sponge.

   For small-scale incision sites cotton-tipped applicators piece of work all-time. Culling scrubs such every bit chlorhexidine may as well be used

4. Echo the alternating scrubs at least iii times.

   Be careful not to excessively wet the creature equally this can exacerbate hypothermia.

Performing Surgery

The pre-surgical preparation should have included consideration of the surgical technique that will be used: sterile surgical gloves (Basic Protocol ii) or clean exam gloves (Alternate Basic Protocol two) with a "tips-simply" technique. Proper surgical attire for both techniques consists of cap, mask, and make clean lab coat.

Basic Protocol ii: Sterile Surgical Gloves

Using sterile surgical gloves allows you to bear upon all areas of the sterile surgical field and surgical instruments with your gloved hand.

Materials

Surgical attire; cap, mask, lab coat

Sterile surgical gloves

Surgical instruments and equipment

Surgical drape

Anesthetized and surgically prepped brute

1. Don cap, mask, and clean lab glaze.

2. Ready the sterile surgical instruments on a sterile surgical field.

   If using surgical packs, verify that the sterilization indicator has turned the appropriate color before using. Unproblematic-pare packs are opened in a fashion that preserves the sterility of the inside surface. Do non touch the within surface as information technology can be used as a sterile field on which to keep the instruments. Circuitous surgical packs are also opened in such a fashion as to keep the inside surface of the wrapping sterile so that it can be used as a sterile field. Instruments in common cold sterilant solutions must be removed from the solution and rinsed with sterile water, saline or alcohol. This is very of import, every bit the sterilization solution is very irritating to tissues. Rinsed instruments must be placed on a sterile field. Dry heat sterilized instrument must also be placed on a sterile surgical field. Remember that bead sterilizers but sterilize the tips of the instruments.

3. Open all other sterile equipment, such equally scalpels and suture material. Open these items in such a way as to preclude contagion of the item and the surgical pack.

4. Place the anesthetized and surgically prepped animal on the warming device that has been covered with a clean paper demote towel.

5. Don the surgical gloves as described beneath (Video 1.12.i) to prevent contamination of the outer surface of the glove (Knecht,1987).

   1. Open the glove packet in such a style that prevents contamination of the inner surface.

   2. With one manus, elevator a glove from the opened packet by its turned-downward cuff.

   3. Pull the glove onto the opposite hand with a rotating motility. Do non affect the exterior surface of the glove.

   4. Place the gloved fingers beneath the cuff of the other glove.

   5. With the gloved fingers under the cuff, pull the glove onto the ungloved manus. The folded cuff protects the gloved hand from contamination.

   6. Pull the cuff of the glove up and over the cuff of the lab glaze.

   7. Slip the fingers nether the cuff of the kickoff glove to pull it over the lab glaze cuff

6. Organize the instruments. It is helpful to point all the tips in one management and place them in the order used (Figure 1.12.11). Between surgeries or during breaks in surgeries cover the tips of the instruments with sterile gauze or drapes (Figure i.12.12).

   

   Organize the surgical instruments.

   

   Between surgeries, the tips of the instruments should be covered.

7. Drape the surgical site.

   The most common mantle is a paper drape ( Figure one.12.13). It may be precut or 1 in which you must cut your ain pigsty. The disadvantage of paper drapes is that they usually cover the entire animal, making patient monitoring hard. Plastic drapes (Figure 1.12.14), usually with an adhesive, offer the advantage of more visibility and better patient monitoring. Sterile gauze sponges (Figure 1.12.xv) tin can likewise be used for drapes. If using the "tips-only" technique you must handle the drape only past its edges then that it does not become contaminated.

   

   

   Plastic agglutinative surgical mantle.

   

   Sterile gauze pads used for surgical drapes.

8. Perform the surgical procedure.

9. Close the surgical wound in layers (e.g., body wall, subcuticular space, peel).

10. Movement animal to a warm cage for recovery.

11. Sterilize or sanitize instruments between surgeries.

    If multiple surgeries are performed each twenty-four hour period and multiple surgical packs are non available, the instruments should exist rinsed with 70% alcohol or "flash" sterilized between surgeries. Remember, alcohol will disinfect, not sterilize. Alternatively, a drinking glass bead sterilizer tin can be used to sterilize the tips of the instruments. Recall to allow instruments to cool before touching tissues!

12. Rinse gloves with 70% alcohol betwixt surgeries.

    If you lot have had to handle another animal to anesthetize and prep it, you should change gloves before performing the next surgery.

Alternate Bones Protocol ii: Clean Test Gloves

Using clean examination gloves and a "tips-but" technique restricts you to using simply the sterile working ends of the surgical instruments to manipulate the surgical field. The gloved, but non sterile, hand must never touch the working end of the instruments, the suture, suture needle, or any part of the surgical field. This technique is useful when working alone and manipulation of not-sterile objects (e.g., anesthesia machines, microscopes, lighting) is required.

Additional Materials (also see Basic Protocol two)

Make clean exam gloves (non sterile surgical gloves)

1. Don cap, mask, and clean lab coat.

2. Place the anesthetized and surgically prepped animal on the warming device that has been covered with a make clean paper demote towel.

3. Don make clean examination gloves.

4. Place the sterile tips of the instruments on a sterile gauze sponge or curtain to prevent contagion (Figure ane.12.sixteen).

   

   Sterile tips of the instruments are placed on a sterile field.

5. Open all other sterile equipment, such as scalpels and suture cloth. Open these items in such a way as to prevent contagion of the item and the sterile tips of the instruments.

6. Drape the surgical site. Handle the drape only by its edges then that it does not become contaminated.

7. Continue as described in steps viii through 12 to a higher place

Commentary

When performing multiple rodent surgeries information technology is a good thought to have staging areas for the different steps of the process. Whenever possible, animals waiting for surgery should be kept at a visual and olfactory distance from those animals undergoing surgery.

As you are performing your surgery, you should be aware of the space that is not sterile between your pack and the draped animal (Effigy ane.12.17). Do not lay instruments in this space. They will become contaminated.

Arrows signal space between drape and instruments that is not sterile.

While performing surgery, be careful not to get paper or fabric instrument drapes wet (Effigy 1.12.18). Wet textile acts as a wick to pull bacteria through from the non-sterile surface below. When this occurs the instruments should be considered contaminated and re-sterilized earlier further use.

A moisture surface area on a newspaper or material pall acts as a wick to pull bacteria through from the non-sterile surface below.

You must go on the recovering patient warm. Do not lay recovering animals directly on the bedding. They may aspirate and asphyxiate. Recovery from anesthesia can besides be aided past the administration of warmed fluids given subcutaneously or intraperitoneally.

In neonates, or animals recovering from prolonged surgical procedures, hypoglycemia can be a trouble leading to postal service-surgical complications. These animals may benefit from the administration of oral glucose. Glucose solutions should never exist given subcutaneously (SQ) or intraperitoneally (IP). Animals may be returned to their holding area once they are awake and appear to be making a normal recovery. Be sure to mark the cage card with the surgical procedure performed and the engagement.

The Guide (NRC, 1996) states that the application of prophylactic antibiotics is not a substitute for the practice of proper aseptic surgery. If prophylactic antibiotics must be used, for example in gastrointestinal surgery or an accidental break in aseptic technique, choose an appropriate antibody and give information technology at the dose and for the length of time recommended by the veterinarian. In guinea pigs and hamsters, the use of inappropriate antibiotics can cause fatalities.

Postal service-operative care does non finish with the return of the animal to its home surroundings. Animals must exist monitored for several days after the surgical process for the evolution of post-surgical complications and for the continued demand for analgesics. Food intake may be difficult to monitor in rodents, particularly if they are group housed. Withal, if postal service-operative animals are singly housed and nutrient rations are supplied in measured amounts this can be a useful monitoring tool. A more applied and sensitive method of monitoring the animal is daily weighing of the animate being. While subtle changes in the animate being'due south activeness or ambition may not be clinically observed, changes in weight will be quickly detected allowing appropriate clinical intervention to exist instituted. It is important to remember that some analgesics may depress the appetite causing secondary weight loss. This weight loss must be differentiated from that which occurs in an animal that is not feeling well.

Supplying a softer, more palatable, easily accessible diet may encourage the animal to eat.

The animate being's hydration tin can be monitored by "tenting" the skin forth the back of the animal. In a well-hydrated animate being, the skin should rapidly fall back into place when released. If an animal is dehydrated, the skin will exist dull to return to its original identify. When this occurs, your veterinary should be consulted for the appropriate utilize of subcutaneous or intraperitoneal fluids.

Wound closures should exist removed at x to 14 days post-operatively. Suture scissors or staple removers should be used.

​

Donning sterile surgical gloves.

Supplementary Material

gloving movie

Video one.12.1:

Donning sterile surgical gloves.

Footnotes

Diskette Information: MAC, Microsoft Word 2004 eleven.2, File proper noun(s): Techniques in Aseptic Rodent Surgery final.doctor, Techniques in Aseptic Rodent Surgery legends and figures.doctor

Publisher'due south Disclaimer: This unit was prepared by Patricia Brown in her private capacity. No official back up or endorsement by the National Institutes of Health or the The states Department of Health and Human Services is intended or should exist inferred.

References Cited

• Flecknell PA. Laboratory Animal Amazement. 2. Academic Press; London: 1996. [Google Scholar]
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• National Research Council. Guide for the Intendance and Use of Laboratory. National Academy Press; Washington, DC: 1996. Veterinary Care; pp. 556–570. http://books.nap.edu/readingroom/books/labrats/ [Google Scholar]
• Swindle MM, Vogler GA, Fulton LK, et al. Preanesthesia, anesthesia, analgesia, and euthanasia. In: Trick JG, Anderson LC, Loew FM, Quimby FW, editors. Laboratory Animal Medicine. Academic Printing; San Diego, CA: 2002. pp. 955–966. [Google Scholar]

Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2587003/

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