Animal-BasedResearch

at theUniversity of Toledo

Lisa J. Root, DVM, MS, DACLAMAttending Veterinarian

Director, Dept. of Laboratory Animal Resources

Overview

1. Animals in Research2. Components of an Animal Care

Program Animal Care (DLAR) Compliance (IACUC) Health & Safety

3. Training4. Occupational Health5. DLAR Vivarium Orientation

Research Animals

The role of animals in medical advancement

CancerDiabetesInfectious diseaseGenetic diseaseOrgan transplantationVeterinary medicine

Animals in Research

Research animals by species: Rodents: 95% Monkeys: 0.5% All others: 4.5%Mice and humans share >95% DNA

http://www.harlan.com/models/athymicnudemouse.asp

Oversight of Animal Research

Laws and RegulationsAnimal Welfare Act and Regulations

All animals* in registered research facilities

USDA Unannounced inspections

annually

*Warm-blooded vertebrates other than mice, rats, and birds

Oversight of Animal Research

Laws and Regulations Public Health Service Policy

All animals* used in research funded by PHS/NIH

The Guide for the Care and Use of Laboratory Animals

Office for Laboratory Animal Welfare

*All vertebrate animals

Accreditation of Research Animal Facilities

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

Non-Profit, Private, Voluntary

AAALAC accreditation demonstrates that the organization meets all regulations and guidelines, and going the extra step to achieve excellence in animal care.

Over 900 institutions in 38 countries are AAALAC-accredited

Program review and site visit every 3 years

UT Animal Care Program

USDA registrationPHS NIH OLAW AssuranceAAALAC Accredited!Health Science Campus has been

AAALAC accredited since 1982.Main Campus joined in 2010.

Animal Care Program

ComponentsInstitutional OfficialLaboratory animal

care (DLAR)Ensures proper care of the

animals

Institutional Animal Care and Use Committee (IACUC)Approves all research

activities involving animalsTraining and Occupational

Health

Health & Safety

IACUCHealth &

Safety

PIs

DLAR

Institutional Official

Lab Animal AidesShirley Jones

Cindy Kokensparger Joyce SweeneyRobert Schuster

Josh Neuman, BS, ALATDustin CoburnJustine Farmer

Nick White

Anna (Jeanna) Ronau Secretary

Frank Calzonetti, PhD Vice President of Research

& Sponsored Programs Institutional Official

President Gaber

Board of Trustees

Department of LaboratoryAnimal Resources

Institutional Animal Care and Use Committee (IACUC)

Scott Bechaz, BS, ILAM, LATgAssociate Director

Lisa Jane Root, DVM, MS, DACLAM Director & Attending Veterinarian

Kathryn Eisenmann, PhDChair, IACUC

Elaine Joseph, PhDResearch Compliance Officer

University of Toledo Animal Care Program

Kimberly Schmidt, BA, LVTAnimal Health and Training Technician

Ashley Kurth, BS, LATgSupervisor

Jamie Harris, LVT, LATgAnimal Technician II

Viviana Ferreira, DVMVice Chair, IACUC

IACUCCompliance

Oversee and enforce

DLAR ≠ IACUCMembership

VeterinariansScientistsNon-scientistsCommunity

members

Review :Animal research

protocolsAnimal Care

Program semiannually

Training and Occupational Health

Investigate concerns

DLARAnimal handlingVeterinary CareSpecific Pathogen Free (SPF)QuarantineTraining

DLAR—Protecting animal health

www.parks.tas.gov.au/.../mammals/rodents.html http://www.radil.missouri.edu/info/dora/Dora.htmwww.usda.gov

UT Animal ResearchHealth Science CampusMain Campus

PsychologyKinesiology

Lake Erie Center

TrainingRequired by USDA

and PHSEnforced by

IACUCPersonnel must

be:Knowledgeable

about the species Fully qualified by

training and experience

ComponentsAnimal handling

and restraint AnesthesiaSurgery Aseptic procedures,

sterile instruments, pain relief, post-operative care

Euthanasia

Controlled Substances and Expired materialsControlled Substances

Drugs that may be misused Regulated by the DEA Can only be obtained by

a DEA license holder

Each PI or department is required to obtain a DEA license

Requirements: Security

Recordkeeping Cannot share controlled

substances with unlicensed individuals or labs

Expired materials

Expired anesthetics, analgesics, and euthanasia drugs can never be used on animals.

Other expired drugs and materials may only be used on animals during a non-survival procedure.

Must be stored separately and clearly labeled as expired.

Occupational Health ProgramHealth Risks When Working With

Research Animal FacilitiesHazards

PhysicalChemicalBiological

AllergiesZoonoses http://www.utoledo.edu/depts/safety/Animal_Research.html

Consideration of Alternatives to Animal Research = 3 R’s

Replacement – non-animal models & animals lower on the phylogenetic scale

Reduction – strategies for obtaining comparable levels of information from the use of fewer animals or maximizing the information obtained from each animal used

Refinement – modification of husbandry or experimental procedures to enhance animal well-being and minimize or eliminate pain and distress

Reporting animal welfare concerns “No facility employee, committee member, or laboratory

personnel shall be discriminated against or subject to any reprisal for reporting violations.”- Animal Welfare Act

To file a concern: Contact a UT official

Provide a description of date, time, location, animal species, numbers and identifications of animals, personnel involved, and any other relevant details

The complainant’s identity will be kept confidential.

Anonymous reports are also accepted if sufficient detail is provided to allow adequate investigation of the allegations.

IACUC Office Dr. Elaine Joseph 419-383-4251IACUC Chair Dr. Kathryn Eisenmann 419-383-4101IACUC Vice Chair Dr. Vivianna Ferreira 419-383-6533

Attending Veterinarian Dr. Lisa Root 419-383-4314

THANK YOU!

Dr. Lisa Root

Lisa.Root@utoledo.edu

419-383-4314 Office in HEB room 012D (in DLAR office suite)

Welcome to Department of Laboratory Animal Resources

“DLAR”

Vivarium Orientation

Lisa Root, DVM, MS, DACLAM

Director

T: 419-383-4310

C: 617-833-3398

E: Lisa.Root@utoledo.edu

DLAR Leadership

Ashley Kurth, BS, LATg Lab Animal SupervisorT: 419-383-2066 E: Ashley.Kurth@utoledo.edu

Jeanna Ronau, BA Office AdminT: 419-383-4310 E: Anna.Ronau@utoledo.edu

Scott Bechaz, GILMA, BS, RLATgAssociate DirectorT: 419-383-5279C: 419-205-0094E: Scott.Bechaz@utoledo.edu

Dr. Lisa Root, DVM, MS, DACLAMDirector & Attending VeterinarianT: 419-383-4310C: 617-833-3398E: Lisa.Root@utoledo.edu

DLAR Technicians

Jamie Harris, LVT, LATg, Lab Animal Technician IIPhone: 419-383-1499 E: Jamie.Harris@utoledo.edu

Shirley Jones, Animal AidE: Shirley.jones@utoledo.edu

Cindy Kokensparger, Animal AidE: Cindy.Kokensparger@utoledo.edu

Josh Neuman, BS, ALAT, Animal Tech1E: joshua.neuman@utoledo.edu

Joyce Sweeney, Animal AidE: Joyce.Sweeney@utoledo.edu

Rob Schuster, Animal AidE: Robert.Schuster@utoledo.edu

Dustin Coburn, BS, Animal AidE: Dustin.Coburn@utoledo.edu

Kimberly Schmidt, BA, LVT, LATAnimal Health and Training TechnicianPhone: 419-383-1499 E: Kimberly.OMearaSchmidt@UToledo.Edu

Justine Farmer, Animal AidE: Justine.Farmer@utoledo.edu

Nick White, Animal AidE: Nicholas.White@utoledo.edu

Vivarium - Main Phone Line (419) 383-4310HSC Vivarium Staffed: M-F 7:30am - 4:00pmMC Vivarium Staffed: M-F 7:30am - ~10:30amWeekends : ~7:30 – 11:30AMHolidays are covered on an abbreviated schedule

DLAR Contact and Hours of Operation

Important Phone Numbers

Safety and Health: Animal Exposure Surveillance Program

• The Public Health Service Policy on Humane Care and Use of Laboratory Animals requires institutions that receive federal funding to provide an occupational health and safety program for employees, visitors and students that work with or have exposure to research animals.

• The University has an Medical Surveillance Program to assess the risk and avoid potential health problems associated with individuals who are involved in the care and use of research animals.

• Multiple sources estimate prevalence of animal allergies in workers range from 5% to 40%

• 15% of the general population is allergic to animals.

• 30% - 50% of those without a previous allergy history will develop an allergy to lab animals while working in that environment.

*DLAM Zoonoses and Lab Animal Allergies – UNC EHS

Symptoms:

Itching, sneezing, watery discharge, congestion, conjunctivitis, asthma, hives/rash, anaphylaxis

Safety and Health: Precautions

• If you are planning on becoming pregnant, are pregnant, ill, or have impaired immunocompetence you should consult a health care professional/physician regarding such conditions and how it pertains to your working with lab animals here at University of Toledo.

• You can also contact the Dept. of Safety and Health who can assist in evaluating your exposure risk and provide case-specific oversight.

Laboratory mice and rats are bred to exclude all zoonotic agents. Therefore, there is limited concern for disease

from these research mice. Historically, rats have been known to carry a bacterium that causes Rat-Bite Fever.

However, these bacteria have not been found in laboratory rats for decades due to the special efforts of

commercial suppliers to eliminate the bacteria from breeding colonies. There is, however, always concern about

secondary infections that can occur with bites and scratches.

• Thus, handle all rodents with care and always wear PPE.

• If you believe you may be developing chronic allergic symptoms from laboratory animal exposure, please notify Environmental Health and Radiation Safety directly.

If bitten or scratched disinfect the wound or site of exposure with disinfection\antibacterial soap.

• Scrub the wound with disinfecting soap. Squeeze puncture wounds if necessary to express blood while disinfecting.

• Medical attention is recommended and a report of the wound is required by filling out an “Injury/Illness Report for Employees and Students:.

• Notify your supervisor

Safety and Health: Animal Bites and Scratches

Safety and Health:Waste Containers

Prohibited in Animal AreasYou are the only person that may use your UT ID for access to the vivarium

(no lending, even to lab mates). • Any visitors must sign in at the front desk (Visitor policy on the IACUC website)

• No open toed shoes (flip flops etc.)

• Long pants are required.

• No food or beverage consumption in the vivarium (procedure rooms or in the animal rooms)

• Avoid scented products (cologne or perfumes) as this can have a negative impact on rodent breeding.

• Speak softly in animal areas—research and breeding animals can be sensitive to sound

• No music allowed. Use headphones & keep one ear available to hear around you.

• Photography and filming is strictly prohibited unless approved by DLAR management

DLAR Access Reminders Access and keys to DLAR will only be granted following completion of all appropriate

training sessions, as well as the Exposure Profile on the Health and Safety website.

Remember to bring your access card and keys to DLAR each time you come down including the weekends and holidays, once obtained. Security and Engineering can’t verify that you are authorized to enter, so they have been given instructions not to let anyone in.

DLAR is a secure place; so please don’t prop doors open or open the door for anyone else.

Smile you’re on camera! Security camera’s are through the facility

Following Traffic Patterns (Flow)Sterile SPF Transition Containment

SPF Transition• May have some minor rodent

viruses/bacteria of no clinical significance

• Clean cages and materials• May be opened in a “hood” or

outside a “hood”

Sterile • Immunocompromised animals highly susceptible to

infections• Keep rodent pathogens out! • Autoclaved sterilized cages, bedding, food, and water • Only opened in a “hood” Specific Pathogen Free a.k.a. SPF • May have some minor rodent viruses/bacteria of no

clinical significance • Clean cages and materials • Only opened in a “hood”

Containment • Has rodent

viruses/bacteria/parasites• Clean cages and materials• No “hood”

Personal Protective Equipment (PPE) For Sterile SPF Room

Hair BonnetSurgical Mask

GlovesDisposable Yellow Gown

Shoe Covers

Disposable Yellow Gown

Hair Bonnet

Shoe Covers

Gloves

Surgical Mask

Personal Protective Equipment (PPE) For SPF Rm/Area

Hair BonnetSurgical Mask

Gloves Disposable Yellow Gown

Disposable Yellow Gown

Gloves

Personal Protective Equipment (PPE) For Surgery Rm/Area

Hair BonnetSurgical Mask

Gloves Disposable Yellow Gown

Disposable Yellow Gown

Hair Bonnet

Gloves

Surgical Mask

Personal Protective Equipment (PPE) ForTransition SPF Rm/Area

Hair BonnetSurgical Mask

*Gloves (don inside the room)Disposable Yellow Gown

Disposable Yellow Gown

Hair Bonnet

*Gloves (don inside the room)

Surgical Mask

*Gloves (Don Inside The Rm.)

Personal Protective Equipment (PPE) For Containment Area

Hair BonnetSurgical Mask

*Gloves (don inside the animal room/s)Disposable Yellow Gown

Shoe Covers

Disposable Yellow Gown

Hair Bonnet

Shoe Covers

N95 Respirator Mask or PAPr

Good Habits to Form: Keeping Your Areas Clean and Safe

• Check that all items are properly labeled:

• Description of what it is• Expiration dates• Any hazards

• Always clean your space:• Make sure you wipe down the

laminar flow hood.• Clean your bench/surface your

working on.

A consistent physical environment and a disease-free environment is essential for good research.

• Temperature: Set point of 72 degrees +/- 2 degrees

• Humidity: Set point of 40%

• Light cycle: Set point of 12 hours on and 12 hours off

• Sanitized or disinfect equipment

• HEPA filter air in cages using ventilated racks

• Micro-Isolator cages with Microbiological filter tops which filter 99.8% of the particulates

• Proper use of PPE is essential

Environmental Conditions and Containment Integrity

SPF/Conventional Rodent Basics

Mouse Information

Physiological Data

• Body Temp: 35.8c -37.4c (96.6f - 99.7f)• Heart Rate 328-780• Respiration: 90-220 p/min• Weight: Newborn 1.0grams, Adult 25-40 grams • Water Consumption: 4-7 ml, or 1.5 ml p/10g body weight per day• Food Consumption: 3-6g, or 1.5 g p/10g body weight per day• Feces: Firm rice size, dark brown in color• Life Span: 1-3 years

Sexing and Breeding

• Sexual Maturity: 40-60 days / 7 – 8 weeks• Estrous Cycle: 4-5 postpartum estrus and within 24 hours after parturition • Gestation: 19-21 days• Litter Size: 6-12• Cannibalism: To reduce cannibalism, do not disturb cage for 7-10 post

birth, feeding the appropriate diet is essential. • Weaning: Standard weaning is 21 days (depending on strain, size of litter

and protocol) animals must not exceed 30 days of age for weaning. • Breeding span: this depends on the strain. Specialized strains may not

breed past 4-6 months. • Two breeding schemes commonly used for mice• Monogamous pairs 1 female and 1 male• Polygamous or harem breeding.

3-4 females (depending on cage size) and 1 male 2-3 females (depending on cage size) and 2 males

Rat Information

Physiological Data

• Body Temp: 35.9c -37.4c (96.6f - 99.5f)• Heart Rate 250-600• Respiration: 66-144 p/min• Weight: Newborn 5.0grams, Adult male 300-500grams , Adult female

200-400grams• Water Consumption: 24-60 ml, or 10-20 ml p/100g body weight per

day• Food Consumption: per day 15-30g, or 5-6g p/100g body weight • Feces: Firm , dark brown, elongated mass with rounded ends • Urine: clear with light yellow tint• Life Span: 2.5-3.5 years

Sexing and Breeding

• Sexual Maturity: 65-110 days / 2.5 – 3.5 months• Estrous Cycle: 4-5 postpartum estrus and within 24 hours after

parturition • Gestation: 20-22 days• Litter Size: 7-11• Cannibalism: Uncommon in rats unless unhealthy. To reduce

cannibalism, do not disturb cage for 7-10 days post birth, and feeding a breeding diet can aid, contact DLAR for assistance with breeding diet in rat rooms. Fostering is also an option, consult DLAR.

• Weaning: Standard weaning is 21 days (depending on strain, size of litter and protocol) animals must not exceed 30 days of age for weaning.

• Breeding span: this depends on strain. Specialized strains may not breed past 4-6 months.

Mouse Handling & Restraint

• Mice can be challenging to grasp as they are quick to avoid handling. • Care should be taken to handle mice appropriately as not harm the mice and to also avoid getting

bitten. • Restraining the mouse can be done by grasping the base of the tail with one hand and with the

other grasp the loose skin behind its neck. Take extra precaution to avoid getting bitten. When you have a firm grasp you may secure the tail in the same hand you have the scruff in to accomplish a one handed restraint or use the other hand for a two handed restraint.

• Care should be taken not to excite any of these animals. Slow deliberate movements will make the job of restraining much easier. Noise should also be kept to a minimum.

• Light restraint is sometimes best for mice (strain dependent).

Rat Handling & Restraint.

• Rats are docile and easier to handle than mice. • Body Scoop

• Supporting the abdomen area carefully scoop the rat. Support the hind legs with the other hand and transfer the rat to a towel or your sleeved arm.

• Tail holding should only be used for aggressive rats (which is rare). • Grasp the rat by the base of the tail but you must transfer the rat quickly to a towel or

the top of the wire for proper restraint. • Always support the body of the rat, never let them hang from the tail beyond the time

to move them from one location to another. • Restraint may be achieved by creating a harness over the back and shoulders with your

fingers with one hand, then supporting the bottom half of the rat with the opposite hand.

Rat Restraint1. Basic/Four finger hold. Lay hand on rat’s back, place head

between index and middle finger, as close to the base of the fingers as possible. Use thumb and last 2 fingers along body to support chest. Grasp tail base with other hand and support body (Figures 10 & 11).

2. Shoulder/Two finger hold. Grasp the tail and gently pull the rat backwards. With the other hand slide thumb and index finger up under shoulders forcing them upwards until the forelegs are crossed. Backward tension must be maintained on the tail, gently stretching the animal to maintain upward lift of the shoulders. In this position the rat is unable to bite the handler (Figure 12).

3. Combination/Three finger hold. Grasp the tail and while exerting gentle backward tension on the tail, slide the other hand forward, placing the thumb under the leg and shoulder on one side, and the index finger on top of the shoulder and alongside the neck on the other side.

4. The middle finger is place under the foreleg with the index finger sitting above. This hold is only applicable for techniques such as ear tag/notch, where the rat can remain on the work surface. (Figure 13).

5. Towel roll. Place the rat on a hand towel or surgical towel. Drape one end of the towel over the rat and gently roll the rat in the towel securely. Place the rat’s head and upper body under your arm while supporting the lower body with the same forearm or hand. Rats usually feel calm and secure with this hold.

Injection Techniques General Overview

Sterile syringes and needles must be used for all injections. The one time use of disposable supplies insures aseptic techniquesand sharp needles. Always select the smallest gauge needle possible to limit tissue trauma and injection discomfort. A 25-30 gauge needle is recommended, ½” to 5/8” length. Before injecting, check for correct placement by pulling back on the plunger of the syringe to create a vacuum. This is known as aspiration.

SUBCUTANEOUS (SQ) INJECTION

MOUSE: The mouse should be restrained in the normal manner. With your fingers, lift the skin to make a “tent”. Insert needle into the subcutaneous tissue. Aspirate prior to making the injection. Proper placement should yield no aspirate. Inject.

RAT: The rat may be wrapped in a towel for restraint. A small opening in the towel over the dorsal shoulder should be made. With your fingers, lift the skin to make a “tent”. Insert needle into the subcutaneous tissue. Aspirate prior to making the injection. Proper placement should yield no aspirate. Inject.

Injection Techniques General Overview

Sterile syringes and needles must be used for all injections. The one time use of disposable supplies insures aseptic techniques and sharp needles. Always select the smallest gauge needle possible to limit tissue trauma and injection discomfort. A 25-30 gauge needle is recommended, ½” to 5/8” length. Before injecting, check for correct placement by pulling back on the plunger of the syringe to create a vacuum. This is known as aspiration.

INTRAPERITONEAL (IP) INJECTION

MOUSE: Restrain the mouse and tilt so that the head is facing downward and its abdomen is exposed. Insert the needle cranially into the abdomen at a 30-45 degree angle caudal to the umbilicus and lateral to the midline. Aspirate: if nothing is aspirated, inject. If anybody fluid/material is aspirated, the drug in the syringe is contaminated and must be discarded. If greenish-brown fluid is aspirated, this indicates needle penetration into the intestines. This is a health concern—discontinue the procedure and contact the veterinarian. If yellow fluid appears, this indicates needle penetration into the urinary bladder. The animal should remain healthy, and the procedure may be repeated with a new syringe and needle. The recommended needle size for IP injections in the mouse is 25-27 gauge, ½ to 5/8” length.

RAT: This may be done as a two-person procedure where one person restrains the rat and the other injects. Alternatively, the rat may be wrapped in a towel for restraint. Restrain the rat and tilt so that the head is facing downward and its abdomen is exposed through an opening in the towel. Insert the needle cranially into the abdomen at a 30-45 degree angle caudal to the umbilicus and lateral to the midline. Aspirate: if nothing is aspirated, inject. If anybody fluid/material is aspirated, the drug in the syringe is contaminated and must be discarded. If greenish-brown fluid is aspirated, this indicates needle penetration into the intestines. This is a health concern—discontinue the procedure and contact the veterinarian. If yellow fluid appears, this indicates needle penetration into the urinary bladder. The animal should remain healthy, and the procedure may be repeated with a new syringe and needle. The recommended needle size for IP injections is 25-27 gauge, ½ to 5/8” length.

On Line Video Training https://www.jove.com/science-education-library/23/lab-animal-research

On Line Video Training https://www.jove.com/science-education-library/23/lab-animal-research

On Line Video Training https://www.jove.com/science-education-library/23/lab-animal-research

Grimace Scale/Pain Management

Painful procedures• Unless the contrary is established, procedures that

cause pain or distress in humans may cause pain or distress in an animal.

• (US Gov’t Principles for the Utilization & Care of Vertebrate Animals Used in Testing, Research, & Training)

• Sedatives, analgesics and anesthetics must be usedunless it is justified for scientific reasons to withhold them.

• Animal Welfare Regulations

CO2 Euthanasia Stations Your protocol must include

descriptions of approved means of primary and secondary euthanasia.

If you choose to euthanize, please follow the SOP for use which is posted at the euthanasia station.

Mouse cages take ~5 minutes

Rat cages takes ~8 minutes

“Take-out” containers can be used to transport 1 or 2 mice for euthanasia. If euthanizing the entire cage take the entire home cage.

Neonatal Rodent Euthanasia

THE UNIVERSITY OF TOLEDO INSTITUTIONAL ANIMAL CARE AND USE COMMITTEE

SUBJECT: Euthanasia of Neonatal Rodents DATE: June 13, 2018 ____________________________________________________________________________

Guidelines for Euthanasia of Neonatal Rodents Neonatal rodents (mouse, rat, and hamster pups up to 10 days of age) are extremely resistant to hypoxia resulting in a prolonged time to unconsciousness with CO2 is used as euthanasia. For this reason, the American Veterinary Medical Association (AVMA) does not endorse CO2 overdose as a method of euthanasia for neonatal pups. Acceptable methods include overdose with chemical anesthetics, decapitation with surgical scissors, or cervical dislocation. Fetuses and altricial neonates (less than 5 days old) may be euthanized through rapid freezing in liquid nitrogen.

References 1. AVMA Guidelines for the Euthanasia of Animals, 2013 Edition.

https://www.avma.org/KB/Policies/Documents/euthanasia.pdf2. Pritchett K, et al. Euthanasia of neonatal rats with carbon dioxide. JAALAS, 4823-27, 2009. 3. Guidelines for the Euthanasia of Rodent Fetuses and Neonates, ARAC Guidelines 04/10/13.

http://oacu.od.nih.gov/ARAC/documents/Rodent_Euthanasia_Pup.pdf

Proper Docking of Caging

Look for the red locking indicator. If you see one, your cage is not properly docked.

Proper docking of caging on ventilated racks provide both air and water for each individual cage (IVC).

Make Sure Automatic Watering Valves are Present on Vent Racks

Standard Diet and Experimental Diets

Special Diet / Experimental DietsExperimental Diets • Ordered by investigators or DLAR if requested

• Clearly labeled expiration date on all diet boxes/containers

• Diet cannot be expired • All cages receive special diet stickers located in the

animal room. • Must be approved on the protocol.• Fasting must have a start date/time as well as an end

date/time. • If adequate food and/or water is not present in the cage,

animal care staff will make an effort to contact research staff. If contact cannot be made, animal care staff will provide ad lib standard diet and water. It is important to leave an emergency number. You may place this sticker on the back of the permanent cage card for its temporary use.

• If a food and/or water-restricted cage is outside the indicated start or end date/time, animal care staff will make an effort to contact research staff. If contact cannot be made, animal care staff will provide ad lib standard diet and water.

• Research staff is responsible for monitoring and replenishing special food and/or water as needed.

Fasted CageNO Food from: _____________AM PM

Until _____________________AM PM

Contact #________________________Past the time DLAR calls, charges & may feed

Diet RestrictionMeasured diet. Cage may run out.

Lab signs daily when fed.Contact #________________________No signature DLAR calls, charges and may feed

SPECIAL DIET #_______________Lab feeds diet & ensures it does run out.

Contact #__________________________ (No food, DLAR calls, charges & may feed

Special Water Contains: _________________________

Water bottle, “NO WATER VALVE” Lab is responsible for water.

(If empty, DLAR calls, charge & may water)

Diet Supplements

DietGel® Boost: “for Mouse Cages”High calorie supplement that provides energy for weanlings, post-surgical and debilitated and aging animals. o One Boost container will

provide supplement for up to 3 cages.

o Split the container into 3 parts using the supplied petri dishes.

o Once split place a couple piece of the rodent chow on top of the Boost (push it into the Boost)

o Place petri dish of Boost and diet on the floor of the cages to allow the rodents easy access.

Diet and Water: “for Rat Cages”Must be provided for weanlings, post-surgical and debilitated and aging animals. o You may use paper cups or a

crock.o Place diet into the paper cup or

crock and fill with water. When using a crock with smaller rats make sure to use enough diet to fill the crock.

o Place paper dish or crock of water and diet on the floor of the cages to allow the rodents easy access.

Diet and Water: “for mice unable to have Boost”Must be provided for weanlings, post-surgical, injured or ill and aging animals.

oPlace diet in a petri dish and fill with water to moisten.oPlace petri dish in cage and provide dry diet on the floor.

Cage Density Requirements Mice

Mice will be housed:•Standard housing is 4 adult mice per cage

•Cage change cycle every 14 days.

•Investigators may request 5 per cage in ventilated cages •25% - 30% of these cages will no make the 14 day cage change and require a spot change. This can create a variable in your research).

Mouse breading:No more than 2 adults and 1 litter, regardless of the size of the litter, will be allowed in a 67 square inch (standard static mouse) cage or 75 square inch (standard ventilated mouse) cage.

1. The recommended breeding strategy for mice is monogamous pairs in either a standard ventilated or static mouse cage.

2. Breeding trios (1 male : 2 females) or harem breeding (1 male : 3 females) are allowable in a mouse cage only if all but one of the pregnant females are removed by the lab prior to parturition (birth) such that only one litter of pups and two adults remain in the cage after pups are born.

3. If post-partum estrus is used, the first litter must be weaned by 21 days of age to prevent the presence of two litters in a cage, i.e., no extended weaning is allowed if post-partum estrus is used.

Cage Density Requirements for Rats• Follow the density guidelines listed below. Rats are social animals and must be group housed unless there is scientific reason (i.e. nursing

mother or clinical issues).

• You may breed rats but our standard cage requires you to separate out the male prior to the female giving birth. You can not have more than the mother with her pups in a standard cage.

• Cages identified as exceeding the guidelines will be marked as such by DLAR staff.

• Investigative staff will have 2 days to wean / separate the animals according to their own preferences; after two days DLAR will separate and charge.

• When breeding, no more than 1 adult and 1 litter, regardless of the size of the litter, will be allowed in a 143 square inch (standard rat) cage. No more than 2 adults (1 male: 1 female) and 1 litter, regardless of the size of the litter will be allowed in a 268 square inch (standard guinea pig) cage.

1. The recommended breeding strategy for rats is monogamous pairs in a standard rat cage.

2. If post-partum estrus is used, the first litter must be weaned by 21 days of age to prevent the presence of two litters in a cage, i.e., no extended weaning is allowed if post-partum estrus is used.

Standard ventilated/static RAT cage = 143 in2

Weight (Grams) Floor area required (in2)/animal

Max # of animal in cage

Up to 200g 23 6Up to 300g 29 4Up to 400g 40 3Up to 500g 60 2

>500g >70 1

Large static RAT cage = 268 in2

Weight (Grams) Floor area required (in2)/animal

Max # of animal in cage

Up to 200g 23 11Up to 300g 29 9Up to 400g 40 6Up to 500g 60 4

>500g >70 3

Expecting/You’ve got Pups! /Weaning Notice Card

• When an animal is due to have a litter place this card ‘Expecting Pups’ side up to inform any staff of potential pups that could be born.

• At time of delivery turn the card ‘Pups DOB’ side up, placed behind the permanent cage card, and add the ‘Pups DOB’ and ‘Wean by’ date in the spaces provided. Generally, all pups are weaned at 21 days but your wean date may be dependent on the protocol.

• An extended wean date may be needed if pups appear undersized but the female must not have been postpartum bred.

• The back of the Expecting Pups card is used by DLAR in the event a new litter is born on cage change day. We do not want to disturb new mothers with a litter for 3 days.

• Also, on the back side of the card, are two perforated sections that can be torn off when pups are weaned. These two sections are placed on the newly weaned cage to alert DLAR and Research staff that the lixit valve must be activated each day for 7 days to help the newly weaned pups know where to find water.

Front of Card Back of Card

Weaning Requirements for Mice

Successful weaning requires: Accurate sexing of pup. Activating the watering, leaving a drop of

water on the valve for the pups to find. Adding a petri dish of Boost with 2-3

pebbles of food and/or wet food in a petri dish.

Adding food to the bottom of the cage. Checking on the pups 2-3 days after

weaning.

Successful weaning requires: Accurate sexing of pup. Activating the watering, leaving a

drop of water on the valve for the pups to find.

Adding a crock of wet food (powdered or pelleted).

Adding food to the bottom of the cage.

Checking on the pups 2-3 days after weaning.

Weaning Requirements for Rats

Sexing Rodents

Sexing Juvenile mice using the Anal-Genital distance method.

Presence of nipples

Other tips If you are a beginner and unsure of sexing, try this: set up 3 cages; one for the females, one for the males, and one for the babies you are not positive of sex. Once you have separated them into these 3 cages, go back to the ones you are unsure of and look again. If you are not 100% positive of the sex, place him/her into a cage by itself then ask DLAR for help. It is better to ask for help than have accidents happen.

Rodent Identification & GenotypingEar punch - The small amount of tissue collected from 1 or 2 ear punches is adequate for genotyping and may be performed on any age mouse without general anesthesia.

Ear-tagging is a metal tag with an ID# attached to one ear of the animal. Young rats may also be skin tagged on the neck with numbered metal tags. These are usually applied at the time of genotyping or weaning. The disadvantage of ear tagging is that tags may fall out if not properly applied.

Animal Census“Hotel fee for each cage”

Animal Census •Census sheets in each room by PI and protocol #. •Lab may update the census sheet if they are adding/removing cages of animals to/from the room.

Example:June 28th you remove 3 cages from room 50F. Write -3 in the slot for June 28th.

Animal Census“Pup Count”

•It is the Principle Investigator’s/Lab’s responsibility to count the number of offspring in your breeding colony.•Monthly, the number of mice you wean from your breeding colony must be deducted from your approved quantity in Protocol. •Email your count to DLAR front office each and every month.

PIMonth/Year Protocol #

Date # of Pups/Fish123456789

10111213141516171819202122232425262728293031

Total 0

Animal TransfersElectronic Animal Transfer Form on DLAR’s website!

• Animals can only receive procedures that are clearly outlined in your approved protocol.

• You cannot perform procedure(s) on an animal(s) that is not allowed in the specific protocol on that cage card.

• If the procedure you want to do is on a different approved protocol you MUST fill out an Animal Transfer form each time you move animals from one protocol to another.

• Example being a breeding protocol to an experimental protocol or between PIs or protocols.

• Submit electronically to DLAR.• Place a Transfer Sticker on each cage you’re

transferring. • Blank labels are in the DLAR binder in each

animal room or you can pre-print your own labels. DLAR will give you the template.

Protocol #:PI: Dept:

Transfer Date:Phone #

Protocol #: 108648PI: ContiDept: Bio SciencesTransfer Date:Phone #7823

Cage Card Stickers:

B

DLAR Labeling

The Guide states that “social animals should be housed in stable pairs or groups of compatible individuals unless they must be housed alone for experimental reasons or because of social incompatibility…If it is necessary to house animals singly…this arrangement should be for the shortest duration possible.”

The following steps are necessary to comply with this requirement:

1. Justification for singly housing animals must be included in the IACUC protocol. 2. The cage card of an animal that is singly housed must be marked in such a way that the DLAR staff and inspectors can readily

determine the reason for being singly housed. Various colored dots or labels are required to identify the cages and reasons: a. A red dot indicates an aggressive or incompatible animal.

b. A blue dot indicates singly housed for a breeding reason’s (removing the male breeder, removing a pregnant female from harem breeding cage, etc).

c. A green dot indicates an animal that recently had a procedure that requires a healing period.

d. An orange dot indicates a reason other than those noted above. A notation must be made on the cage card indicating the reason for being singly housed.

3. Animals should be returned to social housing as appropriate. Breeding females should be returned to group housing after pups are weaned unless they were bred again. Experimental animals should be returned to group housing upon recovery or completion of the procedure.

4. Incompatible animals should never be group housed. 5. The dots are removable and can be used several times. The labels can be covered with a blank label. Labs must utilize the

dots and labels to identify the cages appropriately. 6. If an animal is singly housed and the cage card is not identified with the

appropriate label, DLAR staff will target the cage with a neon green “Single Housed Animal” card. The Principal Investigator (PI) or their research staff will be required to resolve within 48 hours. Repeated non-compliance practices will be reported to the IACUC.

7. A special diet and/or water, measured food and/or water, fasting food and/or water all require stickers that indicates an animal whose food/water intake is being monitored. See descriptions below.

A

S

RESEARCH STAFF SINGLE HOUSED ANIMAL

Follow Guidelines for Single Housing of Social Animals

SPECIAL DIET #__________________ Lab feeds & ensures diet does NOT run out

Contact #___________________________

No food- DLAR calls, charges & may feed

Diet Restriction labels must be used when a lab will be monitoring food

consumption or when an animal will be fed a certain amount of diet.

Special Diet labels are needed when a diet that DLAR does not supply is being used. The diet number/name

and a contact name must be filled out.

When an animal will be placed on a water bottle, a Special Water label must be placed on the cage with a

description of the contents.

When any animal will be going through a fasting period whether it be for 4 hours or 24 hours, a Fasted Cage label must be

placed on each cage being fasted.

DLAR Cage Card

Blank cage cards are available in all animal rooms for any staff. All spaces should be filled out as completely as possible with the appropriate information. There is also adequate blank space for applying informative

stickers and adding animal identification information.

Breeding Card

These cards should be placed on any cage of animals that will be going through a breeding cycle(s). Each line

should be filled out completely and the chart below filled out as appropriate as litters are born/weaned. This card provides a great advantage in tracking breeding pairs,

cycles, and litter size.

Cage-Based Forms:New Cages/Weans/Separation and Breeding

Protocol #: 108648PI: ContiLab Contact:Contact Phone: #7823

Coral Cage Card

Coral colored cage cards are available for any cage regarded as sterile. These should only be placed on sterile cages. The color alerts staff so that they may

perform proper procedures to maintain sterility for those animals. Immunocompromised animals may require

sterile housing.

Post Procedure Care Card*

In addition to the Protocol #, PI and Ph. #, check the appropriate procedure box and the observation period as defined in the protocol. In the table, enter the date/time,

observation/treatment, and initials. Some common abbreviations are listed above the chart. Any and all

procedures performed on an animal should be notated with a post procedural care card. You may also place a sticker in the table to reference a binder/spreadsheet

for long term observation periods.

Post-Surgical Care Card*

These cards must be posted by the researcher and filled out completely after surgery is performed. The

monitoring and analgesia schedule must be followed accordingly. Some cases may require further

monitoring after the surgery report. The back of the surgical care card can be used to record additional

observations.

*Please give to DLAR clinical staff upon completion of study/treatment/observation period.

ROB Procedure Card*

When performing a retro orbital bleed or injection a procedure card must be filled out completely and placed on the cage card for tracking and monitoring. Animals that have had an ROB or injection performed should be monitored closely to watch

for adverse reactions.

Veterinary Care Card (Front)*Veterinary care cards are used by animal care and research staff to request veterinary evaluation of a potentially ill or injured animal. These should be

filled out as accurately as possible to aid in appropriate treatment. When possible use the supplied rodent diagram to notate the area(s) affected. This is a carbon-based form with two copies. The top copy is removed and placed in the

basket on the door of the clinical team office (093) or can be given to any DLAR staff. The carbon copy (cardboard-like) should be left behind permanent

cage card.**Malocclusion: Malocclusion may be a lifelong condition that requires intensive monitoring and treatment on a weekly basis. Mice with malocclusion should not be bred due to potential hereditary

nature of the condition and euthanasia is recommended. If maintaining the mouse is necessary for your research, the mouse may be maintained for a specified amount of time and DLAR will assess a technical

service charge of $7.50 per treatment (weekly) for the clinical care of the mouse.

Veterinary Care Card (Back)*The back of the veterinary cage card is used to track and document observations and treatment (if prescribed). If warranted by the animal’s condition, euthanasia may result from the evaluation for humane concern. This side should be facing

forward behind the permanent cage card to alert staff to treatment. Separate columns can be used to track treatment and observations of several rodents or

multiple treatments on one rodent.

*Please give to the DLAR clinical staff upon completion of study/treatment/observation period.

Animal Found Dead

When an animal is found deceased staff use this card to alert researchers. There are 3 copies (2 are carbon copies) available and should be filled out

fully and accurately. One copy will be placed on the cage behind the permanent cage card, one copy will be given to DLAR staff, and the final

copy will be attached to the carcass bag which is then placed in the refrigerator for 48 hours.

Cage-Based Forms: Hazardous Agent

• Researcher’s must take Hazard Training class.

• Must fill in the dates and contact information on the approved Hazardous posting.

• Submit Hazard Sign to DLAR at least 5 business days prior to any hazardous use.

• Each cage must have a sticker.

BIOHAZARD CHEMICALStart Date: _________________End Date: __________________Agent: _____________________Route: _____________________

*Gloves (Don Inside The Rm.)

Personal Protective Equipment (PPE) For Hazard Room

Hair BonnetSurgical Mask

*Gloves (don inside the animal room/s)Disposable Yellow Gown

Shoe Covers

Disposable Yellow Gown

Hair Bonnet

Shoe Covers

Surgical Mask

Soiled Caging & Equipment to Soiled Side Cage Wash

Thank you for your time and attention!

We are here to help and assist you with your animal research needs.