PACUC Guidelines Rodent Survival Surgery Guidelines for rodent survival surgery are based on the National Research Council Guide for the Care and Use of Laboratory Animals 8 th edition and the Animal Welfare Act (AWA). These statements apply to survival surgical procedures performed on rodents at Purdue University and the following guidelines are to help investigators in complying with the Animal Welfare Act and the Guide for the Care and Use of Laboratory Animals. Pre- Surgical: 1. A separate facility for rodent surgery is not required but surgery should be conducted in a clean, uncluttered area that is not heavily traveled. This area should be disinfected prior to use and between use (Table 1). The designated surgery area should not be used for any other activities during surgical procedures. 2. A pre-surgical evaluation should be performed to ensure the health of the animal undergoing surgery. 3. Withholding of food is not necessary in rodents unless specifically required by the protocol or surgical procedure. Water should not be withheld unless required by the protocol. 4. Prepare the animal by removing hair from the surgical site. This should be done in an area away from where the actual surgery is to take place. Hair should be removed at least 1 cm from all edges of the surgical site. Hair can be removed by clipping with a #40 clipper blade, shaving with a razor, or using a depilatory cream. All loose hair should then be removed with a vacuum or by some other appropriate method. 5. Clean and disinfect the surgical site with an appropriate skin disinfectant (Table 2). Scrub the area with disinfectant in an increasing circular pattern beginning at the center of the circle (proposed incision site) and working outward. After each scrub, rinse the area using the same circular pattern with either 70% alcohol or sterile water/saline. Repeat this process of scrubbing and rinsing 3 times. Do not over wet the animal as rodents are especially subject to hypothermia. 6. As anesthetized rodents are subject to hypothermia, it is recommended that they be placed on a circulating warm water blanket during preparation for surgery. 7. Ophthalmic ointment must be placed in anesthetized animals eyes to prevent the cornea from drying.
8. Surgeons should wear clean lab coats, mask and sterile gloves. If multiple surgeries are planned, sterile gloves should be worn for each animal and should be changed during the procedure if a non-sterile surface or object is touched. A sterile gown and head cover are recommended for long procedures or major surgeries. Surgical: 1. The sterile surgical field must remain sterile throughout the procedure. 2. All surgical instruments must be sterilized (Table 3) prior to initiating the surgical procedure and must not touch non-sterile areas of the animal or environment during the procedure. If multiple surgeries are to be performed, instruments should be cleaned and disinfected (Table 4) or sterilized between surgeries. A glass-bead sterilizer is available for use from the Laboratory Animal Program for this purpose. 3. Sterile drapes must also be used to maintain a sterile field. 4. Anesthetic depth must be monitored before beginning a surgical procedure to ensure that the animal is adequately anesthetized. 5. During surgical procedures the animals vital signs, i.e. heart rate and respirations, must be monitored. 6. Rodents are subject to hypothermia during anesthesia and surgical procedures. Supplemental warmth can be provided with a warm water blanket and warm sterile fluids (saline or lactated Ringers solution). These fluids can be administered subcutaneously at a rate of 1-2 ml/100 grams of body weight. Care must be taken not to overheat any animal placed on or near a heat source. 7. Surgical incisions must be closed with appropriate materials (Table 5). Post-Surgical: 1. Rodents must be kept warm during the post-surgical recovery period and their vital signs monitored. Care must be taken not to over warm the recovering animal. The animal s temperature should be monitored during this time. 2. Take care that the animal s airway does not become obstructed by bedding or other material during the recovery period. Place the animal on its back or side until the animal starts to move on its own. Not until full recovery should the animal be returned to its normal cage and housing area being careful that any cage mates are not biting or chewing on the animal just recovered. 3. Animals should be monitored at least daily and more frequently if the animal is determined to be in pain. Analgesic medications must be administered according to the specific PACUC approved protocol. 4. If dehydration occurs or an animal has experienced blood loss during surgery warmed fluids should be given beginning at 1-2 ml/ 100 g of body weight subcutaneously. Additional fluids may be needed if significant blood loss occurred or an animal is slow to recover from anesthesia.
5. All procedures, medications, and monitoring must be recorded for each individual animal. 6. Wound clips or external skin sutures should be removed in 10-14 days Definitions: 1. Disinfectant - Disinfectants are substances that destroy or inhibit the growth of microorganisms but do not sterilize the area to which they are applied. 2. Sterilization Is the process of killing all microorganisms present, including spores, by the use of chemical or physical methods. References: 1. Guide for the Care and Use of Laboratory Animals 8 th edition, National Research Council, 2011. 2. Animal Welfare Act Regulations 3. Guidelines for Survival Rodent Surgery http://oacu.od.nih.gov/arac/documents/rodent_surgery.pdf 4. Training in Rodent Survival Surgery CD, ILAR. *For assistance with surgical techniques contact the Training Coordinator @ 494-2521
Table 1 Disinfectants for Equipment and Surface Decontamination Name Example Comments Chlorhexidine Nolvasan, Hibiclens Blood does not interfere with activity. These compounds are effective against many viruses and are bactericidal. Alcohols Quaternary Ammonium Compounds Chlorine 70% Ethanol 70% Isopropanol Roccal, Cetylcide Sodium Hypochlorite (Household Bleach 10% solution Chlorine dioxide (Clidox, Alcide ) Contact time is 15 minutes and longer if the surface is contaminated. Alcohols are not sporicidal. These compounds are rapidly inactivated by organic matter. Not very affective against gram negative bacteria. Good cleaning agents. Corrosive. Can be used as a disinfectant on previously cleaned surfaces. Fast acting and broad spectrum. Use a 1/:10 dilution of bleach with a contact time of 3 minutes to kill vegetative organisms. Follow manufactures label for chlorine dioxide compounds Phenolics Lysol TBQ Have antiviral, antifungal and antibacterial properties. Not sporicidal. Can be irritating. Swine and cats are sensitive to phenols and can be fatal if swallowed or absorbed through the skin. Less affected by organic material. Aldehydes Glutaraldehyde (Cidex ) Disinfectant and chemical sterilant. Bactericidal, fungicidal, sporicidal and virucidal. Good activity in presence of organic matter, noncorrosive to endoscopic equipment, rubber or other plastic equipment. Occupational exposure to aldehydes can have health consequences. Can cause irritation and allergies in susceptible people so skin exposure should be avoided.
Table 2 Disinfectants for Use on the Skin Name Example Comments Chlorhexidine Nolvasan, Hibiclens Active against fungi, bacteria, numerous viruses. Good for use on skin. Alcohol Iodophors 70% Ethyl alcohol 70% Isopropyl alcohol Betadine, use of individual swabs is recommended. Evaporation can contribute to hypothermia in animals. Not a high level disinfectant. Activity is reduced in the presence of organic matter and hard water. Wide range of microbicidal action
Table 3 Instrument Sterilants Method Example Properties Steam sterilization Dry Heat Gas Sterilization Chlorine Plasma Sterilization Autoclave Hot Bead Sterilizer Ethylene Oxide Chlorine Dioxide (Clidox, Alcide ) Sterad Time, pressure, and temperature determine effectiveness. Items should be autoclaved for at least 15 minutes at 121ºC and a pressure of 15 pounds per square inch. Very rapid but only tips of instruments not handles are sterilized. Must cool instruments prior to use. Requires special equipment. Toxic and mutagenic. Must aerate all materials exposed to this compound prior to use. At least 6 hours required for sterilization. Shelf life of solution is 14 days. Organic matter reduces activity. Corrosive to instruments. Sterilization equipment utilizes hydrogen peroxide to sterilize heat sensitive instruments and materials.
Table 4 Instrument Disinfectants Name Example Comments Chlorine Chlorhexidine Alcohol Chlorine dioxide (Clidox, Alcide ) Sodium hypochlorite (Clorox 10% solution) Nolvasan, Hibiclens 70% ethyl alcohol, 70% isopropyl alcohol These agents are corrosive and the presence of organic matter decreases activity. Bacteriocial, virucidal, and fungicidal. Contact time is 3 minutes. Effective against most Gram + organisms and some Gram organisms but not against spores. Chlorhexidines have limited activity against viruses. Contact time is 10 minutes or according to label directions. Broad spectrum activity against bacteria, viruses, and fungi but not against spores. Contact time is 15 minutes.
Table 5 Suture Materials Suture Vicryl, Dexon PDS, Maxon Prolene Silk Stainless Steel Wound Clips and Staples Properties Absorbable 60-90 days Absorbable 6 months Inert material, nonabsorbable Nonabsorbable but may wick bacteria into the wound Nonabsorbable Suture material used for skin closure, in general, must be removed in 10-14 days post surgery. Stainless steel wound clips and staples require a special instrument to remove.