BROOKHAVEN NATIONAL LABORATORY

ANIMAL FACILITY (BLAF)

**POLICIES**

<http://intranet.bnl.gov/medical/BLAF/>

September 2008

**TABLE OF CONTENTS**

**INTRODUCTION** PAGE 1

**POLICIES AND RESPONSIBILITIES**

ANIMAL WELFARE ORGANIZATIONS PAGE 2

INSTITUTIONAL ANIMAL CARE AND USE COMMITTEE PAGE 3

RECORDKEEPING PAGE 4

DRUGS PAGE 4

**BROOKHAVEN LABORATORY ANIMAL FACILITY (BLAF)**

ANIMAL FACILITY ACCESS PAGE 5

EMERGENCY PROCEDURES PAGE 5

ENVIRONMENTAL CONTROLS PAGE 5

TRAFFIC FLOW PAGE 6

**PERSONNEL PROTECTION**

PROTECTIVE CLOTHING PAGE 7

HAZARDOUS MATERIALS PAGE 7

**ORDERING/SHIPPING ANIMALS**

ANIMAL ORDERS PAGE 8

ORDERING ANIMALS FROM NON-APROVED SOURCE PAGE 9

ANIMAL ARRIVAL/QUARANTINE PAGE 9

USER CHARGES PAGE 9

ANIMAL CENSUS and PER DIEM RATES PAGE 10

SHIPMENT OF ANIMALS TO OTHER INSTITUTIONS PAGE 10

**CARE OF ANIMALS**

ANIMAL IDENTIFICATION PAGE 11

ANIMAL HUSBANDRY PAGE 11

ANIMAL FEED PAGE 11

ANIMAL CAGE POPULATION PAGE 12

ANIMAL ROOM MAINTENANCE PAGE 12

SICK OR INJURED ANIMALS PAGE 13

MORIBUND ANIMALS PAGE 13

DEAD ANIMALS PAGE 13

ESCAPED ANIMALS PAGE 13

**VETERINARY CARE**

VETERINARY CARE PAGE 14

SENTINEL ANIMAL PROGRAM PAGE 14

SURVIVAL SURGERY PAGE 14

BLAF PROCEDURE ROOMS PAGE 17

ANESTHETICS, ANALGESICS, TRANQUILIZERS PAGE 18

EUTHANASIA PAGE 18

**USE OF ANIMALS**

TRAINING PROGRAM PAGE 19

REMOVAL OF ANIMALS FROM THE BLAF PAGE 19

ANIMAL TRANSPORT POLICY PAGE 20

ANIMAL TRANSPORT VEHICLE PAGE 21

UTILIZATION OF ANIMALS PAGE 22

TRANSFER OF ANIMALS PAGE 22

**APENDIX I**

GUIDELINES FOR SURVIVAL RODENT SURGERY PAGE 23

**INTRODUCTION**

The Brookhaven Laboratory Animal Facility (BLAF) provides facilities,
services, and information to facilitate effective research using
laboratory animals. BLAF is also obligated to ensure that animal care
procedures and animal experimentation comply with federal, state and
local guidelines and regulations.

The BLAF has an approved Animal Welfare Assurance with the Office of
Laboratory Animal Welfare (OLAW), is registered with the US Department
of Agriculture (USDA) and is fully accredited by the Association for
Assessment and Accreditation of Laboratory Animal Care International
(AAALAC). Brookhaven National Laboratory has been fully accredited since
August 3, 1966.

The BLAF adheres to the United States Public Health Service Policy on
the Humane Care and Use of Animals. Consequently, the Guide for the Care
and Use of Laboratory Animal is the standard for our animal care
program. Our commitment to the policy is affirmed by full accreditation
of our facility and animal care programs by AAALAC.

Our activities are subject to unannounced inspections by the USDA under
the Animal Welfare Act, an annual announced inspection by a
representative of the New York State Department of Health and the
possibility of inspection by an official from (OLAW).

If any BNL employee has any question or concerns regarding the care and
treatment of animals in the facility they should contact the Attending
Veterinarian, the Institutional Official or any member of the
Institutional Animal Care and Use Committee (IACUC). All calls will
remain anonymous and individuals will be protected against reprisals.

BLAF Office hours are 8:30 a.m. to 5:00 p.m. Monday to Friday. The
Animal Facility can be reached at (631) 344-3696 or FAX (631) 344-2193.
After hour emergencies should be directed to the Security Division ext.
2238 or 2222.

POLICIES AND RESPONSIBILITIES

**ANIMAL WELFARE ORGANIZATIONS**

**The Animal Welfare Act**

The care and use of laboratory animals, excluding rodents and birds, is
monitored by the USDA under the provisions of the Animal Welfare Act
(AWA). The AWA sets minimal standards for handling, housing, feeding,
and watering of laboratory animals and it also establishes requirements
for basic level of sanitation, ventilation and shelter. The AWA
continues to be revised and amended since its passage in 1966. Our
Institution is visited twice per year by the USDA REAC veterinary
inspector who checks for compliance with the AWA. The AWA also regulates
the use of anesthetic, analgesics, agents of euthanasia, sanitation
practices, experimental protocol and humane treatments of animals,
review of the IACUC species separation, experimental records, heating,
ventilation, air conditioning and light cycles.

**National Institutes of Health**

Institutions using funds provided by agencies of the Public Health
Service (PHS), such as the National Institutes of Health (NIH), to
conduct research on any vertebrate animal must follow additional
regulations. The PHS requires compliance with the *Guide for the Care
and Use of Laboratory Animals* and requires that the institution send an
assurance agreement to the PHS Office of Laboratory Animal Welfare
(OLAW).

BNLhas filed an Assurance with PHS and adheres to all PHS policies
regarding the use of animals for research. Failure of an institution or
by investigators to comply with PHS policy could result in the
termination of research grants and/or loss of eligibility for new
investigator funding from NIH. The activities of the IACUC assure the
PHS that animal care meets the current humane and scientific standards.

**The Guide for the Care and Use of Laboratory Animals**

*The Guide for the Care and Use of Laboratory* *Animals* (The Guide) is
an 80-page booklet prepared by The Institute of Laboratory Animal
Resources (ILAR). The purpose of the Guide is to help institutions
address issues that concern the humane care, use and maintenance of
laboratory animals. The Guide was first published in 1963 and was last
revised in 1996. It provides guidelines that are based on established
scientific principles, expert opinion, and experience with the methods
and practices consistent with humane care and use of laboratory animal.
The Guide outlines the necessary standards for adequate veterinary care,
facility environment and housing requirements, personnel qualifications,
sanitation standards, surgical and post-operative care, acceptable
euthanasia techniques, facility construction design occupational health
and training.

**Association for the Assessment and Accreditation of Laboratory Animal
Care International (AAALAC)**

AAALAC is an international association that provides peer review and
evaluation for laboratory animal care programs. Certification of a
program by AAALAC is accepted by PHS-DHHS-NIH as assurance that an
institution meets current federal policies related to appropriate
laboratory animal care and use. Accreditation is granted on the basis of
evaluation from two sources:

1\. [Annual reports]{.underline} - submitted to the council of the
AAALAC which includes an inventory of animals (by species), a
description of the Institution\'s animal facilities, a description of
the key personnel in its animal care program, and significant changes in
any of these elements from prior periods.

2.  [Site visits]{.underline} - by groups selected from a list of peer
    group professionals approved by AAALAC at least once every three
    years.

**INSTITUTIONAL ANIMAL CARE AND USE COMMITTEE**

The Institutional Animal Care and Use Committee **(**IACUC) is
responsible for the overall program of animal care and use of laboratory
animals at this Institution and for making written recommendations to
the Institutional Official regarding any aspects of the Institution\'s
animal program, facility or personnel training. The IACUC reviews and
approves, requires modification to or withholds approval of those
sections of applications or proposals related to the care and use of
animals as set forth in the PHS Policy. The IACUC notifies investigators
and the Institution in writing of its decision to approve or withhold
approval of those sections of applications or of modifications required
to secure IACUC approval as set forth in the PHS Policy. The committee
is authorized to suspend an activity if the investigator does not comply
with the regulations.

The committee conducts a semi-annual review of the Laboratory\'s program
for the humane care and use of animals. The review consists of
evaluating current procedures, protocols, and practices involving
laboratory animals in research. Appropriate changes will be made based
on the review. The committee surveys the animal facility biannually,
inspecting the physical plant to determine the quality of the animal
care being provided. The committee also inspects all labs where animals
are housed for more than 12 hours, or where animal research is conducted
in order to ensure the lab meets physical requirements and that research
is being conducted in accordance with the approved protocol.

**Review and Approval of Animal Protocol**

All animal studies involving vertebrate animals regardless of the
funding source are reviewed and approved by the IACUC. Investigators
must submit an application to the IACUC representative who will
distribute it to all IACUC members. Each IACUC member will be provided
with all of the applications to be reviewed and may, upon request,
obtain full committee review of any application. IACUC members give
written approval on each submitted protocol by signing the approval
sheets are filed with each individual application. The IACUC notifies
investigators in writing of its decision to approve, modify or withhold
approval of an application. If the decision is to withhold approval of
an application, a written notification and a statement of the reason(s)
will be provided to the investigator.

All IACUC forms are available on the IACUC website -
<http://www.bnl.gov/ora/IACUC.asp>

Instruction on IACUC protocol submission is in the IACUC Standards Based
Management System (SBMS) -
<https://sbms.bnl.gov/sbmsearch/subjarea/149/149_SA.cfm?parentID=149>

**RECORDKEEPING**

Investigators are responsible for maintaining appropriate records. These
include all correspondence with the IACUC, as well as experiment and
surgical records. Surgical records must include the following:

-   Procedures performed including when and by whom.

-   All drugs administered to the animal, noting the dose, time and
    route of administration.

-   Post-operative care including when, how often and by whom.

-   Post-operative analgesia, noting the dose, time and route of
    administration

**DRUGS**

All drugs must be maintained according to CRC Pharmacy policy.
Controlled substances must be stored in a double-locked box and checked
periodically for expiration dates. Ketamine/xylazine mixtures must
comply with CRC Pharmacy policy and may be maintained for a maximum of
28 days. Saline solutions must be labeled with the date when they are
opened and disposed of with 30 days of opening. Mixtures must be labeled
with date mixed.

brookhaven laboratory animal facility

**ANIMAL FACILITY ACCESS**

Animal users requiring access to the animal facility must complete all
training and be listed under an approved IACUC protocol. The PI must
request, in writing, security card access for the individual.

Individuals requesting a key to the animal facility rooms will be
required to fill out a Key Request Form. All keys must be returned to
the BLAF Manager when they are no longer needed. Keys may not be
transferred from one individual to another without permission from the
BLAF Manager.

**EMERGENCY PROCEDURES**

In the event of a medical emergency or animal bite, notify the BLAF
Manager immediately at ext. 3620 or 3696. For emergencies that occur
after working hours contact the Security Office at ext. 2238 or 2222.

Emergency and after-hours phone numbers are posted on or near the front
doors to the BLAF.

[**FIRE/AMBULANCE** **2222**]{.underline} Call this number for fire and
medical emergencies. If you should discover a fire, sound the alarm by
activating a pull station and then call the fire department from a safe
location. Call this number for medical emergencies, which involve life
threatening injuries such as severe bleeding, an unconscious person, a
person who has stopped breathing or major broken bones.

[**POLICE** **2222**]{.underline} Call this number if you have witnessed
a crime or need a police officer.

####### ENVIRONMENTAL CONTROLS

Ventilation rates are periodically measured using appropriate airflow
measuring equipment. Humidity and temperature are measured directly by
reading instruments located in each animal room and recorded daily by
animal care personnel. Humidity and temperatures are maintained for each
species as specified by the AWA.

Fluorescent lights on a time controlled system provide a diurnal
lighting cycle (12 hours light, 12 hours dark) which provide an
intensity of 30 to 40 foot candles at approximately 3 feet above the
floor. Special lighting such as reverse light cycles must be cleared a
minimum of one week in advance through the BLAF Manager.

**TRAFFIC FLOW**

BLAF operates under a clean -- dirty corridor system of movement to
minimize the spread of pathogens. Please remember not to enter the
"clean" side of the facility. Supplies may be requested as needed from
the veterinary staff.

PERSONNEL PROTECTION

**PROTECTIVE CLOTHING**

All researchers entering animal rooms are required to wear clean, cloth
or disposable lab coats and should wear disposable gloves when handling
animals. Hand soap is available in each animal room and gloves should be
changed between cage handling. Disposable gloves, shoe covers, lab
coats, face masks and head covers are also available. N95 respirators or
higher must be worn by BLAF personnel when working in rodent rooms and
are available for researchers as well. If additional garments are
required please notify the BLAF Manager.

**HAZARDOUS MATERIALS**

**Radioactive Material**

Experiments involving the use of radioisotopes must be approved by the
IACUC and discussed with the Radiation Control Division prior to
implementation. Animal cages, excreta, carcasses and all other
radioisotope contaminated material must be monitored and disposed of in
the proper manner by the investigative staff. The Radiation Control
Division has standard operating procedures outlining proper disposal. A
specifically designated area will be maintained to house the animals
involved and a Radiation Work Permit is required.

**Biohazardous Material**

Experiments involving the use of hazardous agents (i.e. infectious
agents, carcinogens or toxic chemicals) must have prior approval from
the IACUC and must be discussed with the Safety office before
implementation. Experimental Safety Reviews outlined by the Safety
office will be instituted to provide adequate safeguards against the
introduction of potential biohazards agents or substances into the
colony population or surrounding areas.

**Transplantable Tumors**

Samples of all transplantable animal tumors must be tested for the
presence of murine viruses prior to injection. All samples sent for
testing will be at the expense of the investigator. The time period to
test these samples is 28 days from receipt of the materials.
Investigators requiring the use of this service should contact the BLAF
Manager.

######### 

ORDERING/SHIPPING ANIMALS

######### 

######### ANIMAL ORDERS

All laboratory animal procurements must be ordered through the BLAF.
Orders should be received no later than noon the Friday before animal
delivery. Telephone orders will be accepted in emergencies only.
Retrieve animal requisition online at:
<http://intranet.bnl.gov/medical/BLAF/> and send to <desmond@bnl.gov>
for approval.

The approved requisition will be forwarded to the BLAF Manager for
review, approval, and ordering. Animals purchased from **approved**
vendors will be placed in clean animal rooms upon arrival. Non-rodents
should be allowed to acclimate for at least five days before
investigators start them on their research projects. For rodents, at
least twenty-four hours is required before placement on study. This
procedure provides time for the animals to acclimate and reduces
shipping stress.

Most animal orders are one-time deliveries. If it is necessary to
receive an order weekly/monthly then an animal standing order may be
placed. When requesting a standing order, indicate the frequency of
order (weekly, monthly, etc.). Orders may be periodically reviewed for
changes in quantity and/or frequency.

In the event that an investigator must change or cancel an animal order,
they should notify the BLAF as soon as possible. If standing orders are
cancelled temporarily, investigators must indicate the date on which the
order is to be resumed.

All animals are procured through reputable commercial breeders utilizing
conventional or modified barrier facilities, which are extensively
monitored for the presence of pathogens.

Species selection is based upon investigator protocol and supporting
scientific literature. The BLAF Manager in consult with the Attending
Veterinarian, must authorize importation or purchase of animals from
non-approved sources. Other research institutions are considered
non-approved importation sources. All animals from non-approved sources
will be quarantined prior to release into the existing animal colonies.
If any organisms hazardous to humans or animals are found, these animals
will not be released into the current pathogen free colonies. Details
explaining the quarantine procedures are outlined under **ANIMAL
ARRIVAL/ QUARANTINE**.

**ORDER OF ANIMALS FROM NON-APPROVED SOURCES**

Investigators requesting the shipment of animals to BLAF from
non-approved sources, i.e. other research institutions or non-approved
vendors, must consult with the BLAF Manager a minimum of three weeks
prior to shipment. The following information is needed:

Facility name, contact person, expected number of animals, the shipping
facility veterinarians name and phone numbers. Health reports will be
requested and animal health status and quarantine space availability
will determine when these animals may be received. Animals will be
placed into the proper quarantine room (depending on health status) and
additional quarantine procedures may be required. Sentinel animals will
be ordered at the expense of the investigator prior to animal receipt.
The submission of surplus cohort animals supplied by the shipper is
greatly appreciated and can effect a more timely evaluation of the
health status of the animals.

**ANIMAL ARRIVAL/QUARANTINE**

Animals purchased from **approved** vendors will be placed in quarantine
or clean holding areas. If requested by the P.I. newly received rodents,
from the same approved vendors, may be housed in the same room with
their other study animals. No serology is necessary.

Animals that are received from **non-approved** sources (i.e. primarily
other research Institutions) must have a health surveillance report and
colony history submitted prior to acceptance that states that they are
specific pathogen free (SPF). These animals will be housed in a
quarantine room for a minimum of 28 days. Sentinel animals will be
placed in cages in the same room with the newly arrived animals at the
expense of the investigator. Diagnostics are sent for analysis at the
cost of the investigator. Once animals have tested negative for murine
viruses, bacteria, and ecto/endo parasites, they can be introduced into
the primary colony.

Serology, parasitology, microbiology, and pathology of quarantine
sentinel animals will be performed after 21 and 28 days. Animals will
not be moved into the general population until a satisfactory health
report has been received. When quarantine animals have been cleared of
all infectious organisms and internal and external parasites they will
be introduced into the general population. If, however, any of these
animals test positive for any pathogenic organisms additional procedures
as required by the Attending Veterinarian will be followed until the
animals are cleared for release to the general colony.

**USER CHARGES**

All animals are maintained at a per diem rate, which can be obtained
from the Medical Department's Administration or online through the
"Budget Office".

These charges cover routine animal housing, sanitation, health care,
identification, feeding, watering, and monitoring daily, including
weekends and holidays.

**ANIMAL CENSUS AND PER DIEM RATES**

A daily animal census will be maintained by the BLAF staff for each
investigator for each species. The census sheets are located in each
animal room on a clipboard. One sheet will be provided for each
investigator in each room. The staff does a physical count of all
animals in each animal room once a week and records deaths,
terminations, and new arrivals.

Investigators must subtract the animals on the census sheet when
permanently removing animals from the room, otherwise they will be
charged for those animals until the next physical count is performed.

Census sheets are collected on the last day of each month and replaced
by a new sheet.

**SHIPMENT OF ANIMALS FROM THE BLAF TO OTHER INSTITUTIONS**

Investigators requesting shipment of animals to other institutions from
the BLAF should submit a request to the BLAF Manager at least two weeks
in advance and must submit a completed shipping request form:
<http://intranet.bnl.gov/medical/BLAF/Forms.asp>

Animals are shipped Mondays, Tuesdays and Wednesdays. Animals should not
be shipped on Thursdays or Fridays unless absolutely necessary. Advance
notification allows for the health certificates and serology report to
be faxed to the receiving institution, assembly of containers and
notification to the shipping companies. Receiving institutions will
notify us when they can accept these animals into their facilities.
Investigators should keep in mind that international shipments require
extra time to coordinate due to regulations and time restraints. The
BLAF has shipping containers and gel packs available.

care of animals

**ANIMAL IDENTIFICATION**

On arrival, all animals received are placed in appropriate sized cages
and an animal identification card is issued. Cage cards are not to be
removed or defaced in any manner. The following information will be
included on all animal cage cards:

Investigator name, protocol number, extension number, animal strain,
species, age or weight, sex, name of vendor, and date received.

If investigators must separate animals for any reason, the information
on the original cage cards must be duplicated to the new cage cards.
Cage cards are also available in the animal receiving room, 11- 120. All
animals must be identified for proper record keeping and to aid in
locating animals. Methods of identification for species are listed
below:

Rats and Mice ear notch, tail tattoo, ear tag

Baboon tattoo, wall chart

Rabbits ear tag, tattoo

Additional color coding will be used to identify different mouse
genotypes used within an experiment.

**ANIMAL HUSBANDRY**

All animal food, bedding, caging, shipping materials (gel packs,
dividers), and other equipment must be pre-approved by the BLAF Manager.
Approved items must be shipped directly from the vendor.

**ANIMAL FEED**

Animals are fed commercial animal diets manufactured by Ralston Purina.

The food supply includes the following animal diets:

High Fiber Rabbit #5326 Rabbits

> Laboratory Rodent Chow #5001 Rodents
>
> Autoclavable Rodent Chow #5010 Immunocompromised rodents
>
> Jumbo Monkey Chow #5037 Baboons
>
> Mouse Chow #5015 Mice
>
> Pico Mouse Chow #5058 Mice
>
> Pico Rodent #5053 Rat

If investigators require a special feed formula they should consult the
BLAF Manager. All exceptions to The Guide for food/water provision must
be approved by the IACUC.

In some instances (ie. post-operative rodents, spinal cord injury
models, stunted weanlings) it is beneficial to provide food and/or Napa
nectar™ (water replacement) inside the cage instead of in the wire bar
cage lid to insure adequate access. The need for this will be determined
by the BLAF staff, under guidance from the Attending Veterinarian and in
consultation with the Principal Investigator. In addition, the food
pellets may be moistened to enhance palatability or Nutrical gel may be
used. In all cases, the food will be placed inside a container and will
be checked daily. Food containers or Napa nectar™ packs that get grossly
contaminated by feces or urine will be replaced.

**ANIMAL CAGE POPULATION**

Federal guidelines and regulations have been established with regard to
space requirements per animal. Cages containing more than the maximum
number of animals will be divided into additional cages by the BLAF
staff. If experiments require that the cage population must exceed the
recommended numbers of animals, this must be scientifically justified in
the animal protocol and must be approved by the IACUC prior to
implementation. All singly-housed rodents will be provided with
environmental enrichment unless scientifically justified.

**ANIMAL ROOM MAINTENANCE**

The BLAF Staff performs routine animal husbandry procedures daily in
every animal room. Food and water is checked daily, including weekends
and holidays. Animal cages are changed according to the table below.

  ----------------------- -----------------------------------------------
  Mouse cages             Changed once to twice per week

  Rat cages               Changed once to twice per week

  Cage lids               Once every two weeks

  Baboon cages            Every two weeks, pans hosed once to twice daily

  Rabbit cages/pens       Every two weeks, pans two to three times per
                          week/or pens daily

  Cage racks              Once a month

  Feeders and water       Once per week
  bottles                 
  ----------------------- -----------------------------------------------

Rodents are housed on direct bedding with Anderson\'s Bed O\'Cobs, 1/8
inch.

Floors should be swept daily and mopped a minimum of once a week.
Investigators are responsible for cleaning up spills and maintaining
clean work areas in their labs and in their animal rooms. Animal
procedures may not be performed in the animal rooms. Designated areas of
the facility, such as the surgery room, can be used to perform
procedures. If investigators require additional space they should notify
the BLAF Manager so that time and space requirements may be scheduled.

######### 

######### SICK OR INJURED ANIMALS

The BLAF staff will check for sick or injured animals daily.
Investigators or technicians who find a sick or injured animal are
responsible for reporting this to the BLAF Manager. Signs of sickness
include:

> Weight loss, eye or nasal discharge, abnormal posture, coughing or
> sneezing,
>
> lack of stool or urine, diarrhea, vomiting, swelling or wounds on
> animals
>
> Alopecia
>
> Dull coat
>
> Inapetance
>
> Blood in the cage

######### MORIBUND ANIMALS

Principal Investigators and staff must monitor all animals on their
protocols after treatment at least three times during the week to ensure
that they are not moribund. The final authority of whether animals
should be euthanized will remain with Animal Care Personnel and/or the
Attending Veterinarian. Animal Care Personnel will make every effort to
contact the Principal Investigator prior to euthanizing an animal.
Principal Investigators must ensure that Animal Care Personnel have
contact telephone numbers for themselves and/ or designees, including
home telephone numbers and/or beepers.

######### 

######### DEAD ANIMALS

Animal care personnel check for dead animals daily. Unless otherwise
requested all dead animals are placed in the freezer for proper
disposable. The animal death is noted on the cage card, the principal
investigator is informed, and the animal is removed from the census
sheets.

**ESCAPED ANIMALS**

Animals found escaped from their cages while still in their room will be
placed in a cage labeled \"Found Loose\" with the date. If they are not
claimed within 7 days BLAF staff will euthanize the animals. Rodents
found loose outside of any rodent housing area will be euthanized.

VETERINARY CARE

**VETERINARY CARE**

Veterinary care is provided by the Attending Veterinarian, Tom
Zimmerman, D.V.M. His authority and responsibilities include the
following:

-   Authority to suspend an activity involving animals if the activity
    is not being conducted in accordance with applicable provisions of
    the Animal Welfare Act, the Guide, or the Institution\'s assurance

-   Oversight of the animal facility and animal care programs

-   Observation of the animals to assess their health and welfare

-   Instruction on the use of appropriate methods to prevent, control,
    diagnose, and treat disease and other medical disorders

-   Provide guidance to users regarding handling, immobilization,
    anesthesia, analgesia, and euthanasia

-   Monitoring of surgery programs and postsurgical care

-   Review and approval of all research proposals which involve the use
    of laboratory animals

-   Visiting the facility three to four times a month, attend IACUC
    meetings, participate in facility inspections, and provide
    additional consultations as needed

**SENTINEL ANIMAL PROGRAM**

The BLAF staff maintains continuous health surveillance of rodent
colonies through the sentinel program. Most rodent rooms have a sentinel
population that is randomly selected from the native population within
the room. All sentinel animals are exposed to the environment of the
other animals within the room by mixing of dirty cage bedding weekly at
cage changing. The sentinel animal population is screened a minimum of
biannually by a diagnostic laboratory for the presence of specific
murine viruses, specific bacterial agents, and ecto/endo-parasites.

**SURVIVAL** **SURGERY**

**Scope:** These guidelines apply to all surgical procedures performed
on rodents at BNL in which the animals are expected to recover from
anesthesia. Prior to performing any survival surgery techniques on
rodents, an approved IACUC protocol must be in place with appropriately
trained personnel and procedures available.

**General:**

The following principles described in the Guide for the Care and Use of
Laboratory Animals apply to rodent surgery.

1.  • Appropriate pre-operative and post-operative care of animals in
    accordance with established veterinary medical and nursing practices
    are required.

    • A dedicated surgical facility is not required.

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

The Guide states that it is important for research personnel to be
appropriately qualified and trained in all procedures to ensure that
good surgical technique is practiced.

Good technique includes:

1.  • Asepsis,

    • Gentle tissue handling,

    • Minimal dissection of tissue,

    • Appropriate use of instruments,

    • Effective homeostasis, and

    • Correct use of suture materials and patterns.

**Intra-operative and Anesthesia Records**

*The Guide for the Care and Use of Laboratory Animals* and the
*Institutional Animal Care and Use Committee Guidebook* require that
animals under anesthesia be carefully monitored to ensure adequate depth
of anesthesia, animal homeostasis, timely attention to problems, and
support during anesthetic recovery. Monitoring includes, but is not
limited to, checking anesthetic depth and physiological parameters
(minimum: heart rate and respiratory rate) on a regular basis (minimum
every 15 minutes).

Record keeping is an essential component of peri-operative care. For all
surgical procedures, an intra-operative anesthetic record must be kept
and included with the surgeon's report as part of the animal's records.
In addition to the above requirements, the record should include all
drugs administered to the animal, noting the dose, time, and route of
administration. These records should be available to the IACUC and BLAF
staff and any other personnel providing post-operative care.

The required monitoring will vary according to the species and the
complexity of the procedure, but should include:

> ●Adequate monitoring of anesthetic depth and homeostasis
>
> ●Support such as fluid supplementation, external heat, or ventilation
>
> ●Monitoring and support during anesthetic recovery
>
> ●Post-operative monitoring

The following are suggestions from the American College of Veterinary
Anesthesiology for monitoring anesthetized animals:

[Circulation]{.underline}: to ensure that blood flow to the tissues is
adequate.

Methods: Heart rate, palpation of peripheral pulses, ECG, auscultation
of heartbeat, non-invasive or invasive blood pressure monitoring.

[Oxygenation]{.underline}: to ensure adequate oxygen concentration in
the animal's arterial blood.

Methods: observation of mucous membranes color and CRT (should be less
than 2 sec if circulation is adequate), pulse oximetry, blood gas
analysis

[Ventilation]{.underline}: to ensure that the animal's ventilation is
adequately maintained.

Methods: respiratory rate, observation of thoracic wall movement or
breathing bag movement if animal is spontaneously breathing, ascultation
of breath sounds, respiratory monitor, capnography, blood gas
monitoring.

**Personal Protective Equipment:**

1.  1\. Clean lab coat

    2\. Mask

    3\. Surgical gloves

    4\. Head cover

**Pre-Operative:**

1.  1\. Surgery should be conducted in a disinfected, uncluttered area
    that promotes asepsis during surgery (see Appendix, Table 1). Use
    clean underpads, towels. Replace these materials after each surgery
    session.

    2\. The instruments, sutures, etc. should be placed in a specially
    designed pack or wrapped in drapes or cloths, then steam autoclaved.
    Label autoclaved packs with date of sterilization. Any implants
    should be sterilized; fragile implants may be gas-sterilized or
    soaked in a 2% glutaraldehyde solution (soak for 10 hours) or other
    chemical sterilant (not disinfectant) - rinse off with sterile water
    before implanting.

    3\. Prepare the animal by removing hair from the surgical site.
    Perform this procedure in an area separate from where the surgery is
    to be conducted.

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1.  4\. Prepare the surgical site(s) with an appropriate skin
    disinfectant (see Appendix, Table 2).

    5\. Surgeons should wash and dry their hands before aseptically
    donning sterile surgical gloves.

**Operative:**

1.  1\. The animal must be maintained in a surgical plane of anesthesia
    throughout the procedure.

    2\. Begin surgery with sterile instruments and handle instruments
    aseptically (see Appendix, Table 3).

    3\. When using "tips-only" technique, the sterility of the
    instrument tips must be maintained throughout the procedure*.*

    4\. Instruments and gloves may be used for a series of similar
    surgeries provided they are maintained clean and disinfected between
    animals (see Appendix, Table 4).

    5\. Monitor and/or maintain the animal\'s vital signs.

    6\. Close surgical wounds using appropriate techniques and materials
    (see Appendix, Table 5).

**Tips on Maintaining Asepsis:**

7.  1\. Gloved hands should be held elevated above the waist and should
    touch only the surgical incision and sterile objects, i.e. sterile
    instrument tray, sterile drape.

    2\. Once gloved, do not touch or lean over a non‑sterile area. Do
    not drop your hands to your sides. Do not touch gloves to your skin
    or clothes.

    3\. Always lift an instrument from a sterile pouch or sterile
    surface. Do not drag instruments over the pack/drape edges because
    they can become contaminated.

    4\. Do not allow surgical instruments to fall below the edge of the
    table. If an instrument does fall, the instrument is considered
    unsterile and should not be picked up and reused until resterilized.

    5\. Sterile surfaces are to be kept dry. Moisture can lead to
    contamination of the surgical area.

**Post-Operative:**

1.  1\. Move the animal to a warm, dry area and monitor it during
    recovery. Return the animal to its routine housing only after it has
    fully recovered from anesthesia.

    2\. Provide analgesics as appropriate and approved in your IACUC
    protocol.

    3\. Generally, remove skin closures 10 to 14 days post-operatively.

    4\. Maintain a surgical record as described above.

5\. Check animals at least daily for the first 3 days after surgery and
make daily notations in the record about the animal's disposition and if
additional analgesia or other treatment is provided.

**BLAF PROCEDURE ROOMS**

The BLAF has procedure rooms available for investigators conducting
animal manipulations, surgical procedures and/or routine handling and
sampling of their research animals. These rooms may be reserved in
advance (on a first-come, first-served basis) by notifying the BLAF
staff and by signing the room usage schedule located in the individual
laboratory. The following guidelines must be followed when using any
BLAF procedure room. Any exception to this policy must receive prior
approval from the BLAF Manager.

1.  Equipment, monitors, or devices may be placed in the room with prior
    approval of the BLAF Manager.

2.  All equipment must be on readily mobile devices.

3.  No fixed equipment or shelving shall be placed in any animal
    procedure room.

4.  All equipment must be removed from the procedure room within
    twenty-four hours if asked to do so by the BLAF management.

5.  Stainless steel surgery or support tables may be placed in procedure
    room with the permission of management.

6.  All needles for injection of substances used on a routine basis are
    discarded in the red biohazard containers. Disposal of sharps or
    syringes in any other container is prohibited.

7.  Broken glass must be cleaned up and disposed of in designated
    containers located in room 11-105.

8.  Eating and drinking in an animal procedure room is prohibited by
    Federal Regulations.

9.  All dirty cages, bottles, and accessories must not remain in the
    procedures room. All items are to be removed and brought to room
    11-118 for cleaning.

######## ANESTHETICS, ANALGESIC AND TRANQUILIZERS

Federal law requires that anesthetic or analgesic drugs appropriate to
the species and procedures in question be used in animal experiments
involving pain or distress.

Investigators are responsible for obtaining all drugs required for
experimental purposes. The BLAF maintains a small amount of drugs for
veterinary health emergencies that are kept in a locked box in an area
of the animal facility.

All isoflurane machines must be calibrated on a yearly basis.

**EUTHANASIA**

The BLAF staff is authorized to euthanize animals. Investigators may
perform animal euthanasia only after receiving appropriate training and
authorization by the Attending Veterinarian. The American Veterinary
Medical Association, Panel on Veterinary Euthanasia is the recognized
authority for all animal euthanasia performed at the BNL. A euthanasia
request form must be filled out prior to the BLAF staff performing
euthanasia. This form can be obtained online at the following address:
<http://intranet.bnl.gov/medical/BLAF/>

**Species Acceptable methods Special Consideration**

Rodents Carbon dioxide 100% with 20% air replacement

CO2/Cervical dislocation Training/proficiency required

Decapitation Scientific justification required

IACUC approval required

Training/proficiency required

> Rats below 200 grams only
>
> Barbiturates

Lagomorphs Barbiturates

Primates Pentobarbital

USE OF ANIMALS

**TRAINING PROGRAM**

The *Guide for the Care and Use of Laboratory Animals*, and the Public
Health Service Policy require that all individuals who are required to
work with laboratory animals in biomedical research be trained to the
rules and regulations governing animal use. These programs require
training in the humane methods of animal experimentation, proper care
and handling of animal species and appropriate use of anesthetics,
analgesics, and tranquilizers.

A training program is available to all BNL investigators, students and
technicians who will have direct contact with the laboratory animals.
Individual or small group training sessions are provided as needed to
help familiarize participants with aspects of the laboratory animal
research program. Sessions are tailored to the individual needs of the
researchers, and consist of hands-on instruction and web-based training.
Additional training seminars are available to research staff members
throughout the year.

**REMOVAL OF ANIMALS FROM THE BLAF**

Animals may be taken from the BLAF only with advance notification to the
IACUC for approved animal workspace and authorization from the BLAF
Manager.

When animals are removed from the facility they must be signed out on
the "Animal Tracking Log" that is located in individual animal rooms.

Animals must be carried in appropriate cages covered by microisolator or
filter top lids, with adequate food and water. In addition, the cages
should be covered so that the animals are not visible. Cover pads and
carts are available for transporting animal cages to and from
laboratories. All caging materials and animal carcasses must be returned
to the BLAF.

When returning empty cages, place them in the dirty side cage-washing
room 11-105 on a cart, if it is available. If animals must be
transported to other buildings on-site, approved vehicles must be used.

Federal requirements state that animals must not be housed outside an
approved animal housing facility for more than 24 hours (12 hours for
USDA-covered species), but in rare circumstances, animals may need to be
housed outside the BLAF for more than 24 hours.  For short term housing,
prior approval by the BLAF Manager is required and the space must be
inspected and approved by the BLAF Manager and/or IACUC.  The full IACUC
must approve housing for periods of longer than one week. Please
complete the following steps:

-   Complete the Non-BLAF Animal Housing Request and submit it to the
    BLAF Manager or designee at least three days prior to the
    anticipated start date;

-   Use the Animal Husbandry Log Sheet to document required checks such
    as lighting, temperature/humidity, food and water;

-   Update the BLAF Manager twice a week with the status of the animals
    and maintain the Lab Daily Activity Record available for inspection.

-   All forms may be obtained on the BLAF website:
    <http://intranet.bnl.gov/medical/BLAF/Forms.asp>

# ANIMAL TRANSPORT POLICY

1\) Drivers must be BNL (or DOE) employees or guests with a valid
driver\'s license appropriate for the operation of the vehicle.
Occupants in government vehicles must be BNL (or DOE) employees or
guests on official business.

2\) Animals shall be transported in air-conditioned/heated vehicles
[only]{.underline}. Auxiliary ventilation such as fans, blowers, or air
conditioners shall be used in the event the ambient temperature ranges
(below) are exceeded during transport.

3\) Overcrowding of the animals' primary enclosure is unacceptable.
Vehicles transporting animals must be large enough to accommodate the
shipment. Primary enclosures shall be positioned in the vehicle in such
a manner that in an emergency the animals can be removed quickly.

4\) During surface transportation the ambient temperature inside a
primary conveyance used to transport animals must be maintained as
follows:

**Nonhuman primates, {between 45^o^ F (7.2^o^ C) and 85^o^ F (29.5^o^
C)**

**Rabbits, Rodents {between 45^o^ F (7.2^o^ C) and 75^o^ F (23.9^o^ C)**

5\) The animal cargo space of the transport vehicle shall be constructed
and maintained in a manner to prevent the ingress of engine exhaust
fumes and gases during transport of animals.

6\) Live animals shall not be transported with any material, substance
(e.g., dry ice), or any device, which may cause injury of the animals.

7\) The interior of the animal cargo space shall be kept clean.

8\) Animal primary enclosures shall be handled with care. (Not tossed or
tipped)

9\) BLAF staff must be notified immediately if there are problems during
transport of animals, or serious unscheduled delays.

NOTE: An approved Radiation Work Permit (RWP) will supercede the
following guidelines if radioactivity is involved.

**ANIMAL TRANSPORT VEHICLE USE, CLEANING, AND MAINTENANCE**

**Primate Transport Vehicle - Chemistry Department**

The primate transport vehicle is typically of a mini-van type, with two
passenger seats and a washable cargo area. The vehicle may be shared
with other department personnel as long as it is cleaned properly before
and after Nonhuman primate use. The researcher responsible for
transporting the animal shall:

1\) Before placing the animal transport cage in the van, apply a heavy
plastic mat, at least four feet in length, over the cargo van floor.

2\) When removing the animal from the van, check the plastic mat for
signs of obvious leaks from the transport cage unit.

3\) If the mat contains spilled material, apply an absorbent product to
the surface immediately.

4\) After using the vehicle to transport animals, the rolled plastic
mat, and/or plastic divider should be removed and cleaned with hot water
and disinfectant.

5\) The floor should be sprayed with disinfectant.

**Animal Transport Vehicle - Medical Department**

This vehicle is of the mini-van type, with two passenger seats and
washable cargo area. It is mainly used to transport small animals
on-site to either the shipping department or NASA holding facility at
the AGS. It can be used to transport different species of animals, but
not at the same time. The researcher responsible for transporting the
animals shall assure that the vehicle is cleaned properly after each
use, particularly between species. Brushes, brooms, and other cleaning
items are available in the BLAF.

**Personal Vehicles**

Cars, trucks, and vans belonging to BNL employees may be used to
transport laboratory animals (rodent species only) with prior
instruction from the BLAF Manager. [At no time will any animal be
transported by bicycle, in the trunk of a car, or in the open cab of a
vehicle.]{.underline}

NOTE: The use of government/contract vehicles is preferred for all
animal transport, and required when transporting non-rodent species or
irradiated animals both onsite at BNL and offsite

**UTILIZATION OF ANIMALS**

The BLAF staff will euthanize animals, which are no longer needed for
research projects. Several programs have been instituted to insure full
utilization and conservation of our animal resources. Some animals that
have been slated for euthanasia will be transferred to the animal
facility census and used as a sentinel animal population for that
particular room. Additionally, animals that are to be terminated may be
used for new investigators, or technician/staff training programs that
require basic hand-on techniques. Animals may also be made available for
collection of blood and/or tissues samples for use by investigators.

Investigators who have made requests for tissues samples will be
notified prior to disposal of these animals.

**TRANSFER OF ANIMALS**

Animals may be transferred from one investigator's protocol to another.
The *Animal Transfer Form* must be completed and approved prior to
animal transfers. *Animal Transfer Forms* can be obtained from the BLAF
staff or online at: <http://intranet.bnl.gov/medical/BLAF/>

All animal records will be evaluated and the appropriate approved
protocols must be on file.

APPENDIX I

**Guidelines for Survival Rodent Surgery**

This appendix includes definitions, tables of information, and
references as a resource for investigators.

DEFINITIONS:

ASEPTIC SURGICAL PROCEDURES: Surgery performed using procedures that
limit microbial contamination so that significant infection or
suppuration does not occur.

MAJOR SURGERY: Any surgical intervention that penetrates and exposes a
body cavity; any procedure that has the potential for producing
permanent physical or physiological impairment; and/or any procedure
associated with orthopedics or extensive tissue dissection or
transection.

MINOR SURGERY: Any surgical intervention that neither penetrates and
exposes a body cavity nor produces permanent impairment of physical or
physiologic function. Examples are superficial vascular cut down, and
percutaneous biopsy.

STERILIZATION: The process whereby all viable microorganisms are
eliminated or destroyed. The criterion of sterilization is the failure
of organisms to grow if a growth supporting medium is supplied.

DISINFECTION: The chemical or physical process that involves the
destruction of pathogenic organisms. All disinfectants are effective
against vegetative forms of organisms, but not necessarily spores.

**Table 1. RECOMMENDED HARD SURFACE DISINFECTANTS** (e.g., table tops,
equipment)

Always follow manufacturer\'s instructions for dilution and expiration
periods.

+-----------------+-------------------+-------------------------------+
| **AGENT**       | **EXAMPLES** \*   | **COMMENTS**                  |
+-----------------+-------------------+-------------------------------+
| Alcohols        | 70% ethyl alcohol | Contact time required is 15   |
|                 |                   | minutes. Contaminated         |
|                 | 85% isopropyl     | surfaces take longer to       |
|                 | alcohol           | disinfect. Remove gross       |
|                 |                   | contamination before using.   |
|                 |                   | Inexpensive.                  |
+-----------------+-------------------+-------------------------------+
| Quaternary      | Roccal®,          | Rapidly inactivated by        |
| Ammonium        | Quatricide®       | organic matter. Compounds may |
|                 |                   | support growth of gram        |
|                 |                   | negative bacteria.            |
+-----------------+-------------------+-------------------------------+
| Chlorine        | Sodium            | Corrosive. Presence of        |
|                 | hypochlorite      | organic matter reduces        |
|                 |                   | activity. Chlorine dioxide    |
|                 | (Clorox ® 10%     | must be fresh; kills          |
|                 | solution)         | vegetative organisms within 3 |
|                 |                   | minutes of contact.           |
|                 | Chlorine dioxide  |                               |
|                 |                   |                               |
|                 | (Clidox®,         |                               |
|                 | Alcide®, MB-10®)  |                               |
+-----------------+-------------------+-------------------------------+
| Glutaraldehydes | Glutaraldehydes   | Rapidly disinfects surfaces.  |
|                 |                   |                               |
|                 | (Cidex®,          |                               |
|                 | Cetylcide*®,*     |                               |
|                 | Cide Wipes®)      |                               |
+-----------------+-------------------+-------------------------------+
| Phenolics       | Lysol®, TBQ®      | Less affected by organic      |
|                 |                   | material than other           |
|                 |                   | disinfectants.                |
+-----------------+-------------------+-------------------------------+
| Chlorhexidine   | Nolvasan® ,       | Presence of blood does not    |
|                 | Hibiclens®        | interfere with activity.      |
|                 |                   | Rapidly bactericidal and      |
|                 |                   | persistent. Effective against |
|                 |                   | many viruses.                 |
+-----------------+-------------------+-------------------------------+

\* The use of common brand names as examples does not indicate a product
endorsement.

**Table 2. SKIN DISINFECTANTS**

Alternating disinfectants is more effective than using a single agent.
For example, an iodophor scrub can be alternated three times with 70%
alcohol, followed by a final soaking with a disinfectant solution.
Alcohol, by itself, is not an adequate skin disinfectant. The
evaporation of alcohol can induce hypothermia in small animals.

+---------------+----------------+-------------------------------------+
| **AGENT**     | **EXAMPLES**   | **COMMENTS**                        |
|               | \*             |                                     |
+---------------+----------------+-------------------------------------+
| Iodophors     | Betadine®,     | Reduced activity in presence of     |
|               | Prepodyne®,    | organic matter. Wide range of       |
|               |                | microbicidal action. Works best in  |
|               | Wescodyne®     | pH 6-7.                             |
+---------------+----------------+-------------------------------------+
| C             | Nolvasan®,     | Presence of blood does not          |
| holorhexidine | Hibiclens®     | interfere with activity. Rapidly    |
|               |                | bactericidal and persistent.        |
|               |                | Effective against many viruses.     |
|               |                | Excellent for use on skin.          |
+---------------+----------------+-------------------------------------+

\* The use of common brand names as examples does not indicate a product
endorsement.

**Table 3. RECOMMENDED INSTRUMENT STERILANTS**

Always follow manufacturer\'s instructions for dilution, exposure times
and expiration periods.

+------------------+------------------+-------------------------------+
| **AGENT**        | **EXAMPLES** \*  | **COMMENTS**                  |
+------------------+------------------+-------------------------------+
| Steam            | Autoclave        | Effectiveness dependent upon  |
| sterilization    |                  | temperature, pressure and     |
| (moist heat)     |                  | time (e.g., 121oC for 15 min. |
|                  |                  | vs 131oC for 3 min).          |
+------------------+------------------+-------------------------------+
| Dry Heat         | Hot Bead         | Fast. Instruments must be     |
|                  | Sterilizer       | cooled before contacting      |
|                  |                  | tissue. *Only tips of         |
|                  | Dry Chamber      | instruments are sterilized    |
|                  |                  | with hot beads.*              |
+------------------+------------------+-------------------------------+
| Gas              | Ethylene Oxide   | Requires 30% or greater       |
| sterilization    |                  | relative humidity for         |
|                  |                  | effectiveness against spores. |
|                  |                  | Gas is irritating to tissue;  |
|                  |                  | all materials require safe    |
|                  |                  | airing time.                  |
+------------------+------------------+-------------------------------+
| Chlorine         | Chlorine Dioxide | Corrosive to instruments.     |
|                  |                  | Instruments must be rinsed    |
|                  |                  | with sterile saline or        |
|                  |                  | sterile water before use.     |
+------------------+------------------+-------------------------------+
| Glutaraldehydes  | Glutaraldehyde   | Several hours required for    |
|                  |                  | sterilization. Corrosive and  |
|                  | (Cidex®,         | irritating. Instruments must  |
|                  | Cetylcide®,      | be rinsed with sterile saline |
|                  | Metricide®)      | or sterile water before use.  |
+------------------+------------------+-------------------------------+
| Hydrogen         | Actril®,         | Several hours required for    |
| peroxide-acetic  | Spor-Klenz®      | sterilization. Corrosive and  |
| acid             |                  | irritating. Instruments must  |
|                  |                  | be rinsed with sterile saline |
|                  |                  | or sterile water before use.  |
+------------------+------------------+-------------------------------+

\* The use of common brand names as examples does not indicate a product
endorsement.

**Table 4. RECOMMENDED INSTRUMENT DISINFECTANTS**

Always follow manufacturer\'s instructions for dilution, exposure times
and expiration periods.

+--------------+----------------+--------------------------------------+
| **AGENT**    | **EXAMPLES**   | **COMMENTS**                         |
|              | \*             |                                      |
+--------------+----------------+--------------------------------------+
| Chlorine     | Sodium         | Corrosive. Presence of organic       |
|              | hypochlorite   | matter reduces activity. Chlorine    |
|              |                | dioxide must be fresh. Kills         |
|              | (Clorox ® 10%  | vegetative organisms within 3 min.   |
|              | solution)      | Corrosive to instruments.            |
|              | Chlorine       | Instruments must be rinsed with      |
|              | dioxide        | sterile saline or sterile water      |
|              |                | before use.                          |
|              | (Clidox®,      |                                      |
|              | Alcide®)       |                                      |
+--------------+----------------+--------------------------------------+
| C            | Nolvasan® ,    | Presence of blood does not interfere |
| hlorhexidine | Hibiclens®     | with activity. Rapidly bactericidal  |
|              |                | and persistent. Effective against    |
|              |                | many viruses. Instruments must be    |
|              |                | rinsed with sterile saline or        |
|              |                | sterile water before use*.*          |
+--------------+----------------+--------------------------------------+

\* The use of common brand names as examples does not indicate a product
endorsement.

**Table 5. WOUND CLOSURE SELECTION**

+-----------------------+----------------------------------------------+
| **MATERIAL\***        | **CHARACTERISTICS AND FREQUENT USES**        |
+-----------------------+----------------------------------------------+
| Polyglactin 910       | Absorbable; 60-90 days. Ligate or suture     |
| (Vicryl®),            | tissues where an absorbable suture is        |
| Polyglycolic acid     | desirable.                                   |
| (Dexon®)              |                                              |
+-----------------------+----------------------------------------------+
| Polydiaxanone (PDS®)  | Absorbable; 6 months. Ligate or suture       |
| or, Polyglyconate     | tissues especially where an absorbable       |
| (Maxon®)              | suture and extended wound support is         |
|                       | desirable                                    |
+-----------------------+----------------------------------------------+
| Polypropylene         | Nonabsorbable. Inert.                        |
| (Prolene®)            |                                              |
+-----------------------+----------------------------------------------+
| Nylon (Ethilon®)      | Nonabsorbable. Inert. General closure.       |
+-----------------------+----------------------------------------------+
| Silk                  | Nonabsorbable. (Caution: Tissue reactive and |
|                       | may wick microorganisms into the wound).     |
|                       |                                              |
|                       | Excellent handling. Preferred for            |
|                       | cardiovascular procedures.                   |
+-----------------------+----------------------------------------------+
| Chromic Gut           | Absorbable. Versatile material.              |
+-----------------------+----------------------------------------------+
| Stainless Steel Wound | Nonabsorbable. Requires instrument for       |
| Clips, Staples        | removal.                                     |
+-----------------------+----------------------------------------------+
| Cyanoacrylate         | Skin glue. For non-tension bearing wounds.   |
|                       |                                              |
| (Vetbond®, Nexaband®) |                                              |
+-----------------------+----------------------------------------------+

\* The use of common brand names as examples does not indicate a product
endorsement.

**Suture gauge selection:** Use the smallest gauge suture material that
will perform adequately.

**Cutting and reverse cutting needles:** Provide edges that will cut
through dense, difficult to penetrate tissue, such as skin.

**Non-cutting, taper point or round needles:** Have no edges to cut
through tissue; used primarily for suturing easily torn tissues such as
peritoneum or intestine.

**Anesthetic Drug Combinations**

In general, by mixing anesthetic and analgesic drugs, the dose required
for each individual drug is reduced, sometimes quite dramatically. Start
at the low end of the dose range listed; you can always give more if
needed!  Drugs not listed below can be mixed using the same concepts,
mix a sedative or hypnotic with an analgesic. Do not mix drugs in the
syringe until you have determined that they are compatible when mixed.
If in doubt administer separately.

Ketamine/Diazepam: Mix drugs 1:1 by volume and administer 0.1 ml/kg IV
for restraint, anesthetic induction or for non-painful procedures. This
gives excellent muscle relaxation, has minimal respiratory or
cardiovascular depression and the animals wake up smoothly and quickly
(within 10-15 min). Visually, these drugs do not appear to mix
completely. When combined and administered as described, the dose is 5
mg/kg ketamine and 0.25 mg/kg diazepam.

Ketamine/Acepromazine: Mix 10 mg acepromazine (1 ml) with 1 g (10 ml)
ketamine and give 0.1-0.3 ml/kg mixture IM or IV (up to 0.6 ml/kg in
rodents and rabbits). Good for restraint, but not for painful
procedures. When combined and administered as described, the dose is
0.09-0.27 mg/kg acepromazine and 9-27 mg/kg ketamine.

Acepromazine/Butorphanol: Mix drugs 1:1 by volume (using 10 mg/ml
butorphanol) and administer at 0.01-0.02 ml/kg IV or IM. Creates a
hypnotic state that is good for restraint and minor procedures that
cause some pain. When combined and administered as described, the dose
is 0.05-0.1 mg/kg butorphanol and 0.05-0.1 mg/kg acepromazine.

Ketamine/Acepromazine/Butorphanol: Mix 10 mg acepromazine (1 ml), 10 mg
butorphanol (1 ml) with 1 g (10 ml) ketamine and give 0.1-0.3 ml/kg of
mixture IM or IV (up to 0.6-0.8 ml/kg in rodents and rabbits). Good for
restraint and moderately painful procedures. More cardiac and
respiratory depression will be seen with this mixture than with ketamine
alone. When combined and administered as described, the dose is 8-25
mg/kg ketamine, 0.08-0.25 mg/kg acepromazine, and 0.08-0.25 mg/kg
butorphanol. For rodents & rabbits, the dose is 50-67 mg/kg ketamine,
0.5-0.7 mg/kg acepromazine, and 0.5-0.7 mg/kg butorphanol.

Ketamine/Xylazine: Good for restraint and painful procedures. Administer
IM, IP, or IV. More cardiac and respiratory depression will be seen with
this mixture than with ketamine alone. Use 100 mg/ml ketamine and 20
mg/ml xylazine to create any of the mixtures listed below.

  ------------- --------------- ------------- ----------------- -------------------
  **Species**   **Recipe by     **Vol to      **Dose**\         **Sedation
                volume**\       give**\       **(per kg body    insufficient?**\
                **(ket:xyl)**   **(ml/kg)**   weight)**         **May redose
                                                                once\...**

  Mouse         mix 2:1         1.5           100 mg Ket + 10   At 1/2 original
                                              mg Xyl            volume

                mix 8:3         2.75          200 mg Ket + 15   *Not recommended*
                                              mg Xyl            

  Rat           mix 3:2         1.25          75 mg Ket + 10 mg At 1/3 original
                                              Xyl               volume

  Rabbit        mix 4:3         0.6           34 mg Ket + 5.2   At original volume
                                              mg Xyl            
  ------------- --------------- ------------- ----------------- -------------------

**Analgesics**

Buprenorphine -- only opioid with long duration of effect in rodents,
0.01-0.05 mg/kg SQ 8-12 hourly for 24-36 hr.

Buprenorphine may be administered orally in jello cube or jelly (with
IACUC approval).

Jello recipe:

1\. Oral dose rate 0.1-0.5 mg/kg.

2\. Figure an average weight for rats in your group.

3\. Determine dosage. Plan on one ml volume of jello per rat.

4\. Plan on making enough jello for 2X daily dosing for up to 4 days.
Strawberry seems to be the favorite. You may need to use partial portion
of jello package and cut water amount.

5\. Follow the jello recipe up to the part where you dissolve the jello
in hot liquid. Reduce the volume of cold water by the volume of buprenex
you will be adding. Add the cold water. Wait until mix is less that 90F
before adding buprenorphine. Pour into ice cube trays @ 4 ml/section.
Freeze.

6\. Dose by giving 1/4 of a 4 ml cube per rat. Place on cage floor on
top of rodent food or petri dish.

NSAIDs -- pediatric syrups palatable to rats.

1\. Ketoprofen injectable- 2.2 mg/kg PO or SQ SID (once daily). Can be
mixed into berry flavored jelly.

2\. Carprofen injectable -- 5 mg/kg S.Q. SID.

3\. Flunixin (Banamine) - 1.1 - 2.5 mg/kg SQ BID.

Note -- animals should be acclimated to oral medications before the
surgery. When added to the drinking water, rats will initially refuse to
drink until they become adjusted to it, which could be disastrous
postoperatively. Similarly, rats require several days to learn to accept
jello cubes.