LAF (HKUST) User Workshop(2021)

Note: This material serves as part of the training program for animalusers of LAF, HKUSTThe rest of the training program includes a tailor-made hands-ontraining section in the facility, a Biosafety course (provided byHSEO) and an LAF User Manual.

Contents:Part 1 Animal EthicsPart 2 Mouse Colony ManagementPart 3 Basic Manual Skills in Mouse HandlingPart 4 Occupational Safety when Working with AnimalsPart 5 Animal Works in Your Research Laboratory

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LAF User Workshop

Part 1

Animal Ethics, Pain and Distress and the 3Rs

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Keystones in Animal EthicsAristotle (384 - 322 BC): • Animals lack reason, human is at the top of the natural world.

Rene Descartes (1596 - 1650)• Only humans are conscious. Animals are living organic creatures, but they are automata,

like mechanical robots.

Jeremy Bentham (1748 - 1832)• The ability to suffer that should be the benchmark of how we treat other beings• If rationality were the criterion, he argued, many humans, including infants and the

disabled, would also have to be treated as though they were things• The question is not Can they reason? nor, Can they talk? but, Can they suffer?

Richard Martin (1754 - 1834)• In 1822, Richard Martin pushed for the first anti-cruelty bill in the UK parliament.

Peter Singer (1946 -)• In his book Animal Liberation (1975), he criticize testing on animals but willing to accept

such testing when there is a clear benefit for medicine. 3

Consensus of animal experiment• It is the ethical obligation of all personnel involved with the use of animals

in research to reduce or eliminate pain and distress in research animals whenever such actions do not interfere with the research objectives.

Keystones in Animal Ethics

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Discomfort:Discomfort is viewed as a mild form of distress.

Stress:Biological responses that an animal exhibits in an attempt to cope with a threat to its homeostasis (Carstens and Moberg2000).

Pain and Distress

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Pain:• is an unpleasant sensory and emotional experience, • is associated with actual or potential tissue damage, • elicits protective motor and vegetative reactions, • results in learned avoidance behavior, and • may modify species specific behavior (Zimmerman, 1986).

Pain and Distress

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Distress:• Distress is a state at which normal biological responses are no longer

sufficient to achieve return to homeostasis.• Normal biological functions may be disrupted as the animal must devote

substantial effort or resources to challenges emanating from the environmental situation.

• Distress may result in disease or pathological changes.

Pain and Distress

Ref: CCAC training module on: pain, distress and endpoints (Canadian Council on Animal Care)7

The 3 Rs of Humane Animal Experimentation

The 3 Rs:• The 3 Rs stand for Reduction, Replacement and Refinement. • In the book The Principles of Humane Experimental Technique, published in

1959 • The authors Russell and Burch proposed that all research using animals

should be evaluated to see if the 3 Rs could be applied.

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The 3 Rs of Humane Animal Experimentation

Replacement • Methods which avoid or replace the use of animals

I. Absolute replacements • The use of an inanimate system as an alternative (mathematical and

computer models)II. Relative replacements• The use of “less sentient” vertebrates or invertebrates, such as

zebrafish, Drosophila and nematode worms

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The 3 Rs of Humane Animal Experimentation

Reduction • Methods which minimize the number of animal used per experiment with

no loss of useful information

• Examples • reducing the number of variables through good experimental design • improved statistical analysis• sharing data and resources • using genetically homogeneous animals (preferably using a number of

distant inbred strains over an outbred strain) • the use of technology, e.g. imaging to enable longitudinal studies in the

same animals

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The 3 Rs of Humane Animal Experimentation

Refinement • Methods which minimize suffering and improve animal welfare• A change in some aspects of the experiment resulting in

• a reduction of pain, stress or distress that animals may experience.

• Examples: • using appropriate anesthetics and analgesics• avoiding stress by training animals to cooperate with procedures• use of non-invasive techniques• use of enrichments that improve living conditions• establish a scientifically sound earliest endpoints for a study that has

the potential to cause pain or distress

Ref:Training module of Canadian Council on Animal CareNational Centre for the Replacement, Refinement & Reduction of Animals in Research

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Local Legislation & Guideline

• The Hong Kong Code of Practice for Care and Use of Animals for Experimental Purposes. (http://ihome.ust.hk/~webacf/files/codeofpractice.pdf)

• All individual researchers must be licensed from the Hong Kong Government Department of Health, under the Animals (Control of Experiments) Ordinance Cap. 340.

• All individual researchers must be in accordance with the Prevention Of Cruelty to Animals Ordinance, Cap. 169.

• All Genetically Modified animals must comply with the Genetically Modified Organisms (Control Of Release) Ordinance, Cap. 607.

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LAF User Workshop

Part 2

Mouse Colony Management

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Colony Management

• Producing animals for experiments

• support experimental colony• maintain genetic purity

• continuing the line

• not to be used in the near future

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Basic needs of Laboratory animals I

Food supply• Ad libitum feeding (i.e. as desired) of diet with the species-specific nutrients

Water supply• Ad libitum supply of clean water

Housing• Enough space and complexity for species-specific activities, e.g.

• nesting materials for nest building• beddings for burrowing• metal grid lid for climbing• toy for playing• huts for further enrichment

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Basic needs of Laboratory animals II

Social interactions• All laboratory animals should not be housed singly in a cage

• exceptions: • specific protocol needs• aggression behavior of individuals (incompatibility of cage mates)• The mouse is the last remaining of a cage

Lighting• Undisturbed light-dark cycle (circadian rhythm)• Species-specific needs of light intensity

• e.g. rodents need low light intensity at day time (note: bright light for safe human activities only )

Temperature• Constant air temperature (around 21°C) 16

Mouse Reproduction Facts IGestation:• 19 - 21 days • 18.5 days in C57BL/6J mice, 20 days in BALB/cJ mice, and 21 days in A/J

mice

Wean age: • 3 - 4 weeks of age• Weaning refers to removing pups from their home cage, i.e. separating

juveniles from the mother• As the starting point, weaning by the week 4 is recommended for strains in

C57BL/6 background, week 3 for most strains in hybrids, BALB/c, C3H/He, CBA, DBA and FVB genetic background.

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Mouse Reproduction Facts IISexually mature:• 5 - 8 weeks of age• Start mating when animals are 6- 8 weeks of age is recommended• Mice bred too early generally produce small litters• Mice bred too late may result in no litter

Reproductive life span:• 7 - 8 months of age• Producing 4 or more litters (e.g. C57BL/6J produces 5.4 litters on average)

Litter size: • 2 - 12 pups• ~3 pups/litter for some 129 substrains; ~6 pups/litter for C57BL/6J; 12 or

more for NOD/ShiLt strains

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Optimizing Breeding Performance

Mate at the optimal age:• 6 - 8 weeks of age is recommended as the starting point

Keep accurate breeding records:Help identifying problem for earlier action

Replace breeders when they: • produce no litter within 60 days of seting mating.• produce no litter within 60 days of their last litter.• produce litters but do not nurse pups for two to three litters.

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Optimizing Breeding Performance

Consult the suppliers for any special breeding characteristic of the strain• Any defective physiology affecting the mating efficiency or reproductive life

span of the strain?• Does the supplier maintain the lines by crossing homozygous animals or

heterozygous animals?• Are you crossing the animals in a different genetic background which may

aversely affect the breeding performance or embryonic survival?

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• Maintain a minimum of six breeding pairs representing two different generations in your colony.

• Do not eliminate one generation until the next one is producing (for a while, you will therefore be maintaining three generations).

• Keep the age range of your breeders between 2 and 8 months old: older mice may not breed reliably.

• Closely monitor breeding performance: if performance declines, promptly take corrective measures.

• Consider backcrossing a strain approximately every 10 generations to prevent substrain divergence.

• Consider cryopreserving a strain in case breeding performance either declines, ceases, or a catastrophic event threatens your colony.

Managing Small Colonies

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Breeding GroupsMating Scheme No. of

malesNo. of females

Benefit Notes

Monogamous (Pair Breeding )

1 1 •Maximizes productivity of a female due to post-partum estrus

•Allows identifying the dam of a litter

•Production rate: 1 litter per month

Trio Breeding (not recommended due to the limited floor area of mouse cages ref)

1 2 •Maximizes productivity of females (post-partum estrus)

•Mothers assist in raising each other’s pups,

•Efficient use of males

•Production rate: 2 litters per month

•Requires extensive handling to avoid overcrowding

•mother of a pups may not be identified.

Harem 1 3-4 •Most efficient use of male•Useful to quickly expand a colony

•Production rate: varies•Requires transferring

mated/pregnant females to separated cages

•Requires intensive handling22

Precaution on breeding/housing practices I

Do not overcrowd a cage• overcrowded cage:

• Rapid consumption of food/water, • Rapid soiling of beddings• Fighting and stress• Unwanted and unidentified mating • Each adult mouse required ~96.7 cm2 when housed in group

IVC: individually ventilated cages Ref: Guide for the Care and Use of Laboratory Animals, 8th edition, NRC23

Precaution on breeding/housing practices II

Housing density• Multiple mice are normally kept in a Holding Cage with the

number specified in the following table• In a Breeding Cage (i.e. for pups delivery), only one mating pair

(1M+1F) with their litter, are housed (not applicable to the small open cages).

IVC: individually ventilated cages Ref: Guide for the Care and Use of Laboratory Animals, 8th edition, NRC

Cage types (floor areas)

Maximum number of adult mice/holdngcage (with no pup delivery)

Large IVC 5

Small IVC 4

Large Open cage 4

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Precaution on breeding/housing practices

Social housing• Animals should not be singly housed, unless with scientific need or other animal

welfare issue (e.g. aggression behavior).

Weanlings must be separated from the parental cage on time (between 3-4 wkold)• Avoid crowding of cages • Avoid unwanted mattings resulting in the production of unnecessary animals. • Survival of the next litter may be compromised by the older litter

Do not co-cage adult males grouping from different cages (or same cage but once separated) • Smell from “strangers” may initiate fights• Exception: young weanlings may be co-cage early without aggression behavior• Aggressive animals should be separated. 25

Precaution on breeding/housing practices(continue)

Determine precisely the mice you need• Avoid over-producing the animals• Identify and terminate animals not to be used in your work as soon as

possible• The more mice you keep, the more complicated is your colony

management

Do not disturb pregnant/early lactating female and her pups• Avoid cage change • Use of sticker notes to remind LAF staff to skip cage changing, if

required.

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Reference and Recommended reading: Breeding Strategies for Maintaining Colonies of Laboratory Mice: A Jackson Laboratory Resource Manual, The Jackson Laboratory

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Mouse Nomenclature

Standardization of nomenclature:• Aids in scientific communication• Link gene to specific alleles and genetic background• Avoid confusion of similar strains or genetic modifications

Creator Lab code:• Identifying a unique allele • Lab code maintained by the international Laboratory Code Registry • You may apply “Hkust” as the Lab code to those transgenic strains

generated in HKUST.(pls see http://ilarlabcode.nas.edu/search_codes_nodep.php?cat=h)

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Mouse Nomenclature

Reference and recommended reading:JAX Full Guidelines: http://www.informatics.jax.org/mgihome/nomen/gene.shtmlNomenclature for mutant alleles generated in ES cell lines by the International Knockout Mouse Consortium (IKMC):http://www.informatics.jax.org/mgihome/nomen/IKMCnomen.shtml 29

LAF User Workshop

Part 3

Basic Manual Skills in Mouse Handling

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Estimating the Age of Pups

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Postnatal day 2

M FM F FM

• Both male and female have protruding external genitalia

• Male: Longer distance between the external genitalia and the anus

• Female adult: presence of vaginal opening (orifice)• Female adult: present of a furless line between the

vaginal opening and anus• Female pups: pale nipples in pigmented pups

Sexing Mice

FemaleMale

Adults

Postnatal day 6 Postnatal day 16

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Hold the base of the tailplace it on a surface that it can grasp

Grasp loose skin on the back of the neck

Secure the tail with the fourth and fifth fingers

R hand

RL hand L

Restraining a Mouse

Recommended training videos: Mouse restrain: https://www.youtube.com/watch?v=ZBPVvRZvIEw (shared by NIH OACO)Rat restrain: https://www.youtube.com/watch?v=SqGN7TCKYZ8 (shared by NIH OACO)

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Ear punching• Applicable to any mice older than 2wks • Cause less discomfort after procedures (no

bone affected)• Ear disc as biopsy for genotyping• Consult LAF staff for recommendation on ear

marking system

Rodent Identifications

Ear Tagging• Applicable to any mice older than 3wks • No biopsy harvested for genotyping• Special equipment is required

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Toe Clipping• Applicable to the mice at 7 days of age or younger• Applicable only when scientifically justified with no other alternative• Minimize the number of digit clipped• Prefer not to remove toe from fore paws• If forepaw must be used, avoid clipping the hallus (“thumb”) which

affect the grasping ability of the animal. • Small sharp scissors must be used and personnel should be trained in

performing the procedure• Tissue as DNA source for genotyping

Rodent Identifications

Ref: Animal Research Advisory Committee Guidelines, Office of Animal Care and Use, NIHGuide for the Care and Use of Laboratory Animals (NCR 2011)

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Tattoo• Specialized equipment and ink are required• No tissue available for genotyping

Marker pen• Non-toxic, “permanent” marker is required• May last for a couple of days only, remarking

may be required to keep the mark

Rodent Identifications

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Intraperitoneal Injection• Body cavity outside internal organs• At the left lower quadrant of the abdomen, • The needle is pushed at an approximately 10° angle• Insert about 0.5 -1 cm of the needle

Compound Administration

Subcutaneous Injection• Over the shoulder is the most usual site• Grabs loose skin• Insert about 0.5 -1 cm of the needle

Recommended readings and training videos:http://www.procedureswithcare.org.uk/intraperitoneal-injection-in-the-rat/http://www.procedureswithcare.org.uk/intraperitoneal-injection-in-the-mouse/http://www.procedureswithcare.org.uk/subcutaneous-injection-in-the-mouse/

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Intramuscular Injection in Rat• Rat: front or back of the thigh; injection vol. < 0.05ml• Causes pain to the animals due to the distension of

the injected muscle

Compound Administration

Intravenous Injection• Injection site: lateral tail vein• Warming of tail/animals is required to dilate the tail

vein (key of success)• Extensive training is required (Key of success)

Recommended readings and training videos:http://www.procedureswithcare.org.uk/intravenous-injection-in-the-mouse/http://www.procedureswithcare.org.uk/intravenous-injection-in-the-rat/

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General note for material injection• Use a new needle for each animal to reduce risk of infection• Shaving and site disinfection is usually not necessary on rodents• Materials with high and low pH can cause pain on the injection site• Further induction of discomfort can be reduced by injecting material

close to body temperature.

Compound Administration

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Gastric Gavage• Stomach tube or dosing cannulae is

used• Estimate length of insertion externally

before gavage• Extensive training is required• If unintentionally administered into the

lung (indicated by respiratory distress), animal should be euthanized.

Compound Administration

Recommended readings and training videos:http://www.procedureswithcare.org.uk/oral-gavage-in-the-mouse/http://www.procedureswithcare.org.uk/oral-gavage-in-the-rat/

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General Conditions (Guide for the Care and Use of Laboratory Animals, 8th edition, NRC)

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

• At the time of use the aseptic surgery should be conducted in an area which is dedicated to surgery and related activities, and

• at all times during the surgery managed to minimize contamination. • All survival surgery will be performed by using aseptic procedures, including

masks, sterile gloves, sterile instruments, and aseptic techniques.

Survival Surgery

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Important notes to survival surgery• BE FAMILIAR WITH THE SURGERY BY RECEIVING THE APPROPRIATE TRAINING (from LAF staff,

your seniors or experts from another laboratory whoever appropriate)• Avoid using too much disinfectant to prevent hypothermic on the animals• Ointment should be applied to the eye of rodents to prevent drying under

anesthesia• Handling tissues gently with the appropriate dissecting instruments• Keep surgical site minimal• Apply surgical aseptic techniques

Survival Surgery

Recommended readings and training videos:http://www.procedureswithcare.org.uk/aseptic-technique-in-rodent-surgery-tutorial/

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Aseptic techniques • Procedures to eliminate contamination of the surgical site thus wound infection• Wound infection causes

• pain and distress (animal welfare issues) and • physiological changes (affect scientific outcome)

Survival Surgery

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Important notes to aseptic techniques• The appropriate personal protective equipment must be worn.• Wash hand thoroughly to eliminate potential contaminants• Use only sterilized dissecting instrument or tools in contact with the wound• Autoclaved items or purchased pre-sterilized materials may be used.• Alcohol alone is NOT considered as an effective disinfectant• Ideally an independent set of sterilized dissecting instrument should be used for

each animal.

Survival Surgery

Recommended readings and training videos:https://oacu.oir.nih.gov/sites/default/files/uploads/arac-guidelines/rodent_surgery.pdfhttp://www.procedureswithcare.org.uk/aseptic-technique-in-rodent-surgery-tutorial/

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What is euthanasia?• In Greeks eu meaning good and thanatos meaning death. • The term is usually used to ending the life of an individual animal in a way that

minimizes or eliminates pain and distress. A good death is tantamount to the humane termination of an animal’s life.

Important guideline• AVMA Guidelines for the Euthanasia of Animals: 2013 Edition• Euthanasia not accepted by AVMA must be justified by scientific needs

Euthanasia

Recommended reading:AVMA Guidelines for the Euthanasia of Animals: 2013 Edition (see page 48-51 for laboratory animals)

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General considerations• Methods of euthanasia likely to elicit distress vocalizations or pheromones that

other animals in the room could hear or smell. It may be best performed in another location.

• Before euthanasia, minimize activities that contribute to distress including • transport, • handling, • disruption of compatible groups, and • elimination of established scent marks

Euthanasia of Mice and Rats

Recommended reading and videos:AVMA Guidelines for the Euthanasia of Animals: 2013 Edition (see page 48-51 for laboratory animals)

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Noninhaled agent injection

Injection of dissociative agent combinations• Ketamine at lethal does• in combination with xylazine or benzodiazpines (α2-adrenergic receptor

agonist)• via intraperitoneal route

Injection of Barbiturates and barbituric acid derivatives • at three times the anesthetic dose• via intraperitoneal route

Acceptable Method(Mice and Rats)

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Carbon dioxide inhalation• Compress CO2 gas in cylinder as the source, dry ice is NOT ACCEPTABLE• Optimal filling rate to the euthanasia chamber should displace 10-30%

of the chamber volume per minute (strictly follow the specified flow rate post at the euthanasia station in LAF).

• Prefilled chamber is NOT ACCEPTABLE. • Chamber ( if not the home cage) should be emptied and cleaned

between uses.• Death MUST BE confirmed by physical examination, e.g. confirming

breathing and heart beat arrest.• Death may be ensured by a physical method

(e.g. cervical dislocation and decapitation)

Acceptable with Condition Method(Mice and Rats)

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Euthanasia of Rodent fetuses

Fetuses in utero• Fetuses in utero are unconscious• Hypoxia does not evoke a response• Unnecessary to remove fetuses for euthanasia after the dam is euthanized

by acceptable methodFetuses removed from uterus

• Fetuses removed from dams may be aroused• Mouse and rat fetuses of >E15 may be euthanized by decapitation• Fetuses >E15 should be euthanized or anesthetized before chemical fixation• Anesthesia may be induced by hypothermia in cold saline (with the amniotic

sac intact) until the fetuses become immobile

Ref:AVMA Guidelines for the Euthanasia of Animals: 2013 Edition (see page 48-51 for laboratory animals)NIH Guidelines for the Euthanasia of Rodent Fetuses and Neonates

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Euthanasia of Rodent neonates

Acceptable methods for neonates up to 10 days of age • Injection of chemical anesthetics, e.g. pentobarbital, and then decapitation• Anesthetic inhalation, e.g. CO2 or insoflurane, and then decapitation• CO2 inhalation cannot be used alone as the animals are resistant to hypoxia,

decapitation may be use as a secondary physical procedures for euthanasia.

Acceptable methods for neonates over 10 days of age • AVMA Guidelines for the Euthanasia of Animals is applied

Ref:AVMA Guidelines for the Euthanasia of Animals: 2013 Edition (see page 48-51 for laboratory animals)NIH Guidelines for the Euthanasia of Rodent Fetuses and Neonates

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• Let the mouse grasp the grid• Restrain the rodent by grasping the base of

the tail• Place fingers against the back of the neck at

the base of the skull• quickly pull backward with the hand holding

the tail base• verify dislocation by feeling for a separation

of cervical vertebrae • Confirm respiratory arrest, no heart beat

Cervical dislocation

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Common problems in your mouseWhat is abnormal?Observation Possible reason (apart from genetic problem)

Shape Thin Low food consumption, infection/problem with ingestion

Undersize, weak pup Occasional too much competition in uterus/getting milk

Hunched Severe pain/illness due to different reasons

Outgrown teeth Misalign of upper and lower jaw, failed to wear each other

Rectal prolapse Bacterial infection, heavy burden of parasites in guts

Hydrocephalic Cerebrospinal fluid accumulated in the brain, inherited/infection

Movement Spinning/flipping Problem in balance, infection of ears

Paralysis Spinal cord damage, illness

Labored breath Problem with nose/lung, infection, presence of mass/fluid in lungs

Fur/skin Hair loss Behavioral abnormal barbering (one in the cage looks normal), scratching due to parasite/skin infection

Coat color change Genetic contamination

Staining (urine, feces etc) Illness, diarrhea

Wound (tails, hind feet) Fighting, cocaging of non-compatible mice

Scaly skin Infection by Corynebacterium bovis

Ulceration Infection of (fight) wound/tumor outgrowth

Ulceration (Tumor)

Hunched

Hairloss (Barbering)

Rectal prolapse

Hydrocephalic

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LAF User Workshop

Part 4

Occupational Safety when Working with Animals

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Major hazards in working with animals

• Allergens• Zoonotic disease• Protocol related hazards

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Allergy Risk

Facts• 11 and 44% of the individuals working with laboratory animals report work-

related allergic symptoms.

• Of those who become symptomatic, 4 to 22% may eventually develop occupational asthma that can persist even after exposure ceases

Low risk:• Procedures on unconscious animalsHigh risk:• Handling live animals

Ref:Bush and Stave (2003) Laboratory animal allergy: an update. ILAR J;44:28–51

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Major Source of Allergens

Ref:Bush and Stave (2003) Laboratory animal allergy: an update. ILAR J;44:28–51

• Hair, dander, urine and saliva• By contact or respiratory ( on <5micron particles)

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Symptoms of allergy

Contact urticaria• redness, itchiness of skin, welts and hives;Allergic conjunctivitis• sneezing, itchiness, clear nasal drainage, nasal congestion;Allergic rhinitis• sneezing, itchiness, clear nasal drainage, nasal congestion;Asthma• cough, wheezing, chest tightness, shortness of breath;Anaphylaxis• generalized itching, hives, throat tightness, eye or lip swelling, difficulty in

swallowing, hoarseness, shortness of breath, dizziness, fainting, nausea, vomiting, abdominal cramps, diarrhea.

Ref:http://www.iacuc.ucsf.edu/Safe/awSafeOhs.asp

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Minimizing Allergen Exposure

Procedural Control• Replace open-top cages with individually ventilated cages or static cages • Handle animals in biological safety cabinet or animal transfer station• Segregate animal works from other works• Minimizes aeroallergen levels and prevents spread of allergens into the

environment (e.g. wherever appropriate, use female or juvenile animals instead of adult males, as male animals secrete more allergens in their urine)

Personal Control• Correct use of personal protective equipment• General hygiene

Ref:Gordon and Preece (2003) Occupational Medicine 2003;53:371–377

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Response to allergy

• Stop animal works• Report to your supervisor and seek clinical assistance• Ask for clinical consultation on your fitness to continuing animal work

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Zoonotic Disease

Zoonotic disease • Any disease that may be transmitted from an animal to a human under natural

conditions.• Animals used for laboratory research or teaching purposes in LAF include mice,

rats, rabbits and Xenopus.

Transmission of zoonotic disease is primarily by • fecal-oral route, • inhalation of aerosolized materials, • indirect contact with invertebrate vectors and • contaminated inanimate objects.

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Risk of Zoonotic Disease

Imported animals • Laboratory rodents are generally maintained free from specific pathogens. Risk

of zoonotic disease is low.• Agriculture, Fisheries and Conversation Department request imported animals

were maintained in colonies tested free from specified zoonotic pathogens, i.e. hantavirus, lymphocytic choriomeningtis virus and rabies.

Wide rodents• The presence of zoonotic agents are more likelyPet rodents• Pathogens in pet breeding facilities are common • Keeping pet rodents poses high risk to laboratory animals thus the health of

other animal users

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Zoonotic Disease

Prevention of zoonotic diseases• Do not keep pet rodents or snakes which are fed with live rodents• Prevention of bites and scratches,

• Familiarize yourself about the animals that you will be working with• Wear the appropriate gloves

• Post-injury treatment.• Thoroughly wash any bite or scratch wounds and report injuries. Rabies

exposure is reportable to public health authorities• Do not eat, drink and apply makeup while handling animals or in animal housing

areas• Keep animal areas clean and disinfect equipment after using it on animals or in

animal areas

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Protocol related hazards

Biological or chemical hazard• Identify the use of viral vector expressing oncogenic (or suspected oncogenic) or

toxin product, use of infectious agents and tumor cells• Identify the use of carcinogenic or toxic chemicals• Take the respective biosafety/chemical safety precaution procedures (if

required, consult HSEO or LAF for the agent specific-precaution)• Notify LAF for the precaution on the animal and soiled cage handling• Use the appropriate PPE to avoid direct contact with the biological agents• Note personal hygiene to avoid carrying over of the biological agents

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Protocol related hazards

Physical harzard• Flammable substances• Sharps• Electricity shocks by high voltage equipment

The one hand scoop technique

www.gravesassociates.com64

Protocol related hazards

Important notes • be familiar with the corresponding precaution procedures and • seek specific training from LAF or your research group before conducting

procedures involving potential hazard• notify LAF for the proper animals and cage handling

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LAF User Workshop

Part 5

Animal Works in Your Research Laboratory

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Animal Works in Your Research Laboratory

Considerations• Environment in the laboratory is different from a designated animal holding

room.• Inappropriate air temperature control• Lack of tight light dark cycle• High light intensity• Noisy • Not enough fresh air supply to remove allergens

• Inappropriate environment may cause stress to the animals and hazard to laboratory personnel (animal wellbeing issues).

• The increase of frequency and duration of keeping the animal in the laboratory increases the dispersal of allergens to all personnel in the laboratory (occupational safety issue)

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Reason for performing animal procedures in the laboratory

Justify the need of bringing the animals to the laboratory which risks animal welfare and allergen hazard to the laboratory personnel.

• Bringing animal away from the animal facility only when performing a particular procedure in the animal facility is impossible, e.g.• procedures requiring a bulky equipment being shared with other non-animal

users • isolation of tissue must be done shortly before further processing, which

must be performed in the laboratory• Multiple treatment on the animals in a single day, without the need of special

equipment, may be possible in the animal facility.

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Time arrangement on the animal procedures

• The period when the animals staying in the laboratory should be as short as possible as needed. i.e.• prepare the experimental set up and reagents ahead in the laboratory,• remove the animal from the animal holding room immediate before the

procedures and.• return the animals to LAF as soon as the procedures and recovery (if

required) finished.

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Precaution on transferring the animals

• Cages should be covered by the appropriate filter cover to contain allergens and allow the proper air supply

• Avoid bright light by properly shielding the cages• Cages should be carried with a stable trolley/cart or manually• Use air conditioned indoor path whenever possible

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Temporal keeping of animals in the laboratory

• If required, life animals should be temporally kept in a place with the following condition• ambient temperature• quiet • dim • without chemicals around (especially volatile and powder)• NO chemical fume cabinet• NO oven of any temperature for pups (keep pups warm with the original

nest material or finely teased paper tissue)

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Note during the animal procedures

• ALL procedures must be performed as described in an Animal Ethic Committee approved Protocol and by approved persons listed in the Protocol.

• If surgery is performed, the same requirement of aseptic procedures as in PART 3 of this Workshop must be applied.

• If terminal procedure is performed, the animals are euthanized according to the requirement by the AVMA Guidelines for the Euthanasia of Animals (2013 Edition)

• Adult mice are commonly euthanized by cervical dislocation while young pups are euthanized by decapitation.

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Note after procedures

After the procedures• Surviving animals should be returned to the LAF animal holding room 7221 as

soon as the procedure and recovery are completed• Record the procedures and the use of animals accordingly• Cage materials should be doubly wrapped in autoclavable bags and returned to

LAF entrance C as soon as possible.• Animal carcasses should be doubly bagged and return to designated freezers

immediate outside LAF entrances as soon as possible.

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END of LAF User Workshop

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