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CENTRAL LEPROSY TEACHING & RESEARCH INSTITUTE

(DIRECTORATE GENERAL OF HEALTH SERVICE)

MINISTRY OF HEALTH & FAMILY WELFARE

GOVT OF INDIA CHENGALPATTU-603001, TAMIL NADU, INDIA

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CONTRIBUTORS Shri. M. Rajenderen Sr. Technical Assistant

Shri. U. Aravindan Junior Statistical Officer

Dr T. Pugazhenthen Medical Officer

Dr. V. M. Bhagat Assistant Director

Dr. V. C. Giri Deputy Director

Dr. M. K. Showkath Ali Director

PUBLISHER

Central Leprosy Teaching & Research Institute (CLTRI)

Directorate General of Health Service Ministry of Health & Family Welfare, Govt of India

Tirumani, Chengalpattu, Tamil Nadu -603001

Contact us @ dircltri@dataone.in, dircltri.tnchn@nic.in

(044) 27426065, 27426064 Visit us @

www.cltri.gov.in

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TABLE OF CONTENTS Page No

I . THE ENVIRONMENT

A. Climatic control

1. Temperature ……………………………………………………. 04

2. Humidity ………………………………………………………... 05

3. Ventilation………………………………………………………. 05

4. Light……………………………………………………………... 06

B. Other environmental factors

1. Noise……………………………………………………………... 06

2. Odor……………………………………………………………… 06

3. Bedding………………………………………………………….. 07

4. Population Density and Space……..…………………………... 07

C. Minimum Floor Area required for Laboratory Animals…………….

D. Laboratory Animal care 08

1. Well being Laboratory Animals……………………………….. 09

2. Reception………………………………………………………… 09

3. Maintenance…………………………………………………….. 09

4. Identification and Records…………………………………… 10

5. Feed……………………………………………………………… 10

6. Water…………………………………………………………….. 10

II. LABORATORY ANIMAL HANDLING-CARE AND FACILITY 11

A. Cleaning and Sanitation ………………………………………………. 12

B. Wastes Disposal………………………………………………………… 12

C. Vermin Control ……………………………………………………… 13

D. Holiday and Emergency Care…………………………………………. 13

E. Anaesthetic Agents…………………………………………………… 14

III. STANDARD OPERATING PROCEDURES (SOP)………………………...

IV. BASIC DATA: COMMON LABORATORY ANIMALS………………….. 15 16

V. REFERENCES………………………………………………………………….

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Environmental requirements will vary with the species and the

experimental protocol. In general, a constant and comfortable environment

is required to ensure both the health of the experimental laboratory animal

house for meaningful results. Many laboratory animals, especially the

smaller species, establish their own microenvironment within their cages.

The nature of these will vary with the animal’s habits, the type cage and the

climatic conditions prevailing within the room itself.

The animal facility should be designed that the experimental animal

room climate is adjustable to meet the special requirements of the

Investigation and the needs peculiar to the species housed.

To facilitate this it is obviously projects and species and that each

such room have separate environmental controls. In many facilities this

will not seem to be possible through proper planning, colony management

and the installation of automatic light timers, portable humidifiers and air

conditioning units etc.

Most laboratory animals can tolerate the same temperature range as

man, thus the temperature in animal holding rooms tends to be a

compromise between what is best for the animal and most comfortable for

the workers.

1. Temperature

Environment

5

Sudden change in temperature variations may harm laboratory

animals. Emergency equipment to maintain appropriate environmental

temperatures should be available, particularly in buildings where housing

of small laboratory animals, normally the range will be of same as man 200C

to 250C

Most animals prefer a humidity of about 50%, but can tolerate a

range of 30% to 70% as long as the temperature range is appropriate to the

species and the humidity remains relatively constant.

Fluctuations and extremes in relative humidity can precipitate illness,

particularly respiratory diseases.

Dehumidifiers may need to be used where automatic watering and flushing

systems are used in facilities that do not have a controlled environment.

The animal facility should be ventilated properly. It is preferable to

use a total air exchange system. If a recirculation system is to be used, it

should be equipped with effective filters and necessary recirculation of air

should be given careful consideration when planning a new animal facility.

Air conditioning is useful in providing a stable environment of 10-15

changes per hour.

2. Humidity

4. Ventilation

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Light in animal rooms should provide good visibility and uniform,

glare-free illumination. Intensities of between 807-1345 lux at 76cm (30”)

from the floor have been widely recommended to facilitate proper

laboratory animal observation, record keeping and house keeping. Light

intensity in the order of 200 lux has been shown to be adequate for

reproduction and to assure normal social behavior amongst most rodents.

Noise is unavoidable in an animal care facility, but should be

minimized. It can disturb both the animal and staff; unexpected sounds

seem to be more harmful. Loud noises precipitate epileptic form seizures in

several species and strains of animals, intermittent noise may also affect

drug response and breeding performance.

Some animal odors are offensive to humans, and some can

significantly affect the physiological and pharmacological responses in

experimental animals. Much of the odor in an animal facility results from

bacterial decomposition of excreta, and can be controlled by maintaining

the cleanliness and adequate ventilation. Cages as well as the room should

be checked for odors as it is the ammonia (NH3) resulting from the

decomposition of excrement.

5. Light

1. Light

2. Odor

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The choice of bedding materials in case of small rodents, profoundly

affect their microenvironment. In general, small rodents’ longevity tends to

be increase when maintained on bedding like saw powder, paper bits and

paddy husk.

Animals of all species should be provided with solid flooring and

bedding well prior to parturition. Unsterilized saw powder are a possible

source for the introduction of disease, particularly parasites, into the

rodent colony though contamination with cat feces and those of wild

rodents thus affecting the normal physiology of the experimental animal.

The number of animals to be kept to a cage will obviously be

influenced by the demands on existing animal room space, the caging

available, the level of the technician work load and the types of laboratory

animal in use. The details are given below

3. Bedding

4. Population Density & Space

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Animal Weight in gm Floor area/ Animal (cm2)

Cage height (cm2)Polythene/

polypropylene/SS

Mice

<10 Up to 15 Up to 25

<25

38.7 1.6 7.4

96.7

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Rats

<100 Up to 200 Up to 300 Up to 400 Up to 500

>500

109.6 148.3 187.0 258.0 387.0

>=451.5

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Hamster

>60 Up to 80

Up to 100 >100

64.5 83.8

103.2 122.5

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Guinea pigs <350 >350

387.0 >=651.4

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Animals Weight in gm

Floor area (Sq.ft)

Floor area (Sq.mt)

Height (inches)

Rabbits

<2000 Up to 4000 Up to 5400

>5400 Mother with

kids

1.5 3.0 4.0 5.0 4.5

0.135 0.27 0.36 0.45 0.40

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Animals Weight

(Kg)

Floor area

(ft2)

Floor area

(Cm2)

Height

(Cm)

Monkey

Up to 1 Up to 3

Up to 10-12 Up to 12-15 Up to 15-25

1.6 3.0 4.3 6.0 8.0

1440 2700 3870 5400 7200

50 72 72 72 90

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An animal which grows and behaves normally and is free of disease is

usually considered to in a state of “Well-being”. All aspects of animals care

should be directed towards the achievement and preservation of this state.

Its maintenance requires effective health monitoring, suitable exercise. and

Each new shipment of laboratory animals should be received,

examined and placed in clean cages at a quarantine room. Shipping

containers should not enter the main facility and should either be

incinerated, incoming animals should be identified and their arrival

appropriately recorded. The name of the supplier of each shipment should

be noted, along with pertinent observations on the quality and condition of

the animals he has supplied. Animals that appear sick should be euthanized

without delay.

Wherever it is possible species should be housed in separate rooms.

Shipment of the same species, acquired from different suppliers, should

also be separate if space permits. Where the mixing of species and/or

stocks from different sources may be unavoidable every effort should be

made to placed together those that are compatible, have similar

environmental requirements

1. Well-being” of Laboratory Animal

2. Reception

3. Maintenance

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Cage or group identification may be used for small laboratory

animals. Record should include each animal’s arrival time, sex, estimated

age and weight, breed and type, color and marking and any physical

abnormalities or other identifying features. Use of room cards on the doors

of animal rooms indicating the species is a good practice.

All experimental animals should received palatable uncontaminated

and nutritionally-adequate food according to the requirement of the

species. Follow the United Federation of Animal Welfare(UFAW) laboratory

animal feed formulae (Annexure I,II). Whenever possible pasteurized or

sterilized laboratory animal food obtained from standard suppliers should

be used. The storage of bulk feeds should be such as to minimize the

possibility of contamination. Dry pellets stored at room temperature in

cool, dry, well ventilated room. Bulk feeds should not be store in animal

colony. Feed containers should be clean and disinfected frequently.

Slightly acidified drinking water should be available to laboratory

animals at all times, unless contraindicated by the experimental protocol.

Admissible chlorinated watering method unlikely to spread disease or

contaminate the water supply should be chosen.

Water bottles should be clean, clear, transparent, to permit ready

observation of cleanliness and water level. They should be of a material

6. Water

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5. Feed

4. Identification & Records

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that will withstand sterilization and should be of a wide mouth design to

facilitate cleaning water bottle should always be replaced with clean,

freshly-filled Bacterial contamination, particularly with pseudomonas and

coli form organisms can easily occur and must be monitored routinely for

bacterial contaminates.

Laboratory animals may should be handled and restrained when put

into new cages or removed for various experimental purposes. Most

domestic and laboratory animals need no restraint for such routine

handling but will respond to gentleness; in fact they tend to escape from

cages.

Staff should manage the laboratory animals, in order to develop a

sense of security, and learn the minimum amount of force required to

safety hold or restrain various species.

Under normal conditions, all standard laboratory animals, excepting

primates, should be handled with bare hands. In all cases only the minimal

amount of force necessary should be employed. Manipulation of the type

and intensity of light used often proves useful in handling small mammals

and rodents.

Successful handling also requires the ability to recognize the animal’s

state of mind, which may include bewilderment, apprehension and in some

cases discomfort or pain. Whenever possible, the use of gloves should be

avoided, as they often prevent the handler from developing the proper

sense of touch and because the animal should not be discomfort when use

Laboratory Animal Handling

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of productive restring. In this regard, the routine use of specialized cage

squeeze mechanism is advocated for handling the larger non-human

primates.

Cleanliness, including personal cleanliness on the part of the staff,

cannot be overemphasized in an animal care specialty. Employees should

follow the proper cleaning and disinfecting procedures and their

importance in disease prevention.

All cages, pens, racks etc. must be thoroughly cleaned and disinfected

before reuse. As a general rule, the animal house should be cleaned every

day or alternative day. Animal cages are most efficiently cleaned and

sanitized with mechanical washing equipment operating at 83oC (180oF) or

higher, for a minimum of ten minutes.

Cages should be carefully rinsed to remove all traces of washing and

disinfecting agents, Bedding in animals cages or pens should be changed as

often as necessary to keep the animals clean, dry, and relatively odor free.

Smaller laboratory animal require one to three changes per week,

depending on population of the laboratory animals.

Dead animals, animal tissue excreta, bedding, unused diet etc. should

be collected with care an in leak proof metal or plastic containers and

incinerated. Waste which cannot be rapidly disposed of should be stored in

1. Cleaning and sanitation

2. Waste Disposal

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a hold storage area provided for that purpose. Such areas must be vermin-

free, easily cleaned and disinfected as well as been physically separated

from other storage facilities. Dead animals should be properly identified,

placed in disposable plastic bags and immediately incinerated upon

discovery, installing an incineration facility for the disposal of pathological

and animal wastes should be planned for animal house well in advance

during civil and electrical construction

A properly construction building should be vermin proof, but may

not be free from vermin. Vermin enter on feed, bedding, man and

laboratory animals. Insects and arthropods thus introduce may act as the

intermediate hosts of certain parasites and may also mechanically transmit

certain other infections. Wild rodents may transmit a wide variety of

bacteria, viruses, and parasites to the housed species.

New facilities should be checked critically for vermin before any

animals are moved in. training of personnel, good waste disposal, sealing or

eliminating breeding sites, extermination through pesticides or trapping,

and the recovery of all animals. It is important that pesticides should be

applied only under supervision.

Laboratory animal care is a continuous and daily responsibility.

24X365. This point should be emphasized in job descriptions for animal

care personnel as it is most essential service. All the animal care staff

should be informed of other responsibilities in emergency situations.

3. Vermin Control

4. Holiday and Emergency Care

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Drugs

(mg/kg)

Mouse Rat Guinea pig Rabbit Monkey

Ketamine Hcl 22-24 IM 22-24 IM 22-24 IM 22-24 IM 22-24 IM

Pentabarbitone

sodium

35 IV

50 IP

25 IV

50 IP

30 IV

40 IP

30 IV

40 IP 35 IV

Thiopentone

Sodium

25 IV

50 IP

20 IV

40 IP

20 IV

55 IP 20 IV

25 IV

60 IP

Urethane - 0.75 IP 1.5 IP 1.0 IV,IP 1.0 IV

IM- Intramuscular IV – Intravenous IP – Intraperitoneal

5. Anesthetic Agents for Laboratory Animals

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Standard operating procedures

The Institute shall maintain SOPs describing procedures /

methods adapted with regard to Animal Husbandry, maintenance,

breeding, animal house microbial analysis and experimentation

records are follows:

1. Name of the Author

2. Title of the SOP

3. Date of preparation

4. Reference of previous SOP on the same subject and date (Issue

no and Date)

5. Location and distribution of Sops with sign of each recipient

6. Objectives

7. Detailed information of the instruments used in relation with

animals with methodology (Model no., Serial no., Date of

commissioning, etc)

8. The name of the manufacturer of the reagents and the

methodology of the analysis pertaining to animals

9. Normal value of all parameters

10. Hazard identification and risk assessment

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Life Span of Common Laboratory Animals (UFAW)

Approximate daily food intake (UFAW)

Animals Daily Food intake

Mice 5 gm

Rat 15 gm

Monkey 200 gm

Guinea pig 30 gm

Hamster 10 gm

Rabbit 120 gm

Animals Life Span

Rat 2 – 3 years

Mouse 1 – 2 years

Guinea pig 3 – 5 years

Rabbit 5 – 6 years

Monkey 15 – 30 years

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Rat Mouse Guinea

pig

Rabbit Monkey

Age at puberty

(months) 1 – 2 1 2 6 – 9 30 – 36

Minimum

breeding age

(months) 3 1 3 – 4 9 – 12 50 – 56

Oestrus

duration (hours) 10 – 20 10 – 20 6 – 12 Induced 24 – 36

Oestrus

intervals (days) 5 5 14 – 16 None 30

Gestation period

(days) 21 – 22 19 – 21 60 – 80 30 – 32 168

Recurrence

oestrus (months) Post

partum

Post

partum

Post

partum

End of

lactation -

Breeding life of

females (years) 1 1 2 – 3 2 – 3 6 – 8

Breeding life of

males (years) 1.5 1.5 3 1 – 3 8 – 10

Breeding ratio

(M/F) 1:2 1:2 1:5 1:1 1:1

Litter size (Nos.) 5 – 10 7 – 12 2 – 6 4 – 6 1

Birth weight

(grams) 5 – 6 1 – 1.5 50 – 80 80 – 100 300 – 500

Weaning weight

(grams) 35 – 40 10 – 12 250 600 – 800 800 – 1000

Weaning days

(Nos.) 21 19 – 21 21 – 30 45 90 – 150

Breeding data of Laboratory Animals

(CPCSEA guidelines)

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Rat Mouse

Guinea

Pig

Rabbit Monkey

Daily feed

intake (gm) 10 - 15 5 - 10 30 – 50 100 – 200 100 – 300

Daily water

intake

(ml/100gm)

body weight

15 5 – 10 15 20 30 – 40

Urinary output

(ml/100gm)

Body weight 5 – 8 3 – 4 4 – 9 7 – 8 5 – 6

Daily fecal

output (gm) 9 – 13 6 – 9 15 – 18 20 – 30 100 – 150

Pulse rate

(no’s/min) 300 600 150 155 90 – 100

Respiratory

rate (no’s/min) 85 – 113 163 82 – 90 38 – 60 39 – 60

Rectal

temperature (F) 99 – 100 96 – 100 100 – 102 102 – 103 100 – 102

Room

temperature (F) 65 – 75 68 – 74 65 – 75 62 – 68 68 – 72

Relative

humidity of

room (%) 45 – 55 45 – 55 45 – 55 45 – 55 45 – 55

Physiological data of laboratory animals

(CPCSEA guidelines)

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QUANTITY RATS MOUSE GUINEA

PIG

RABBIT MONKEY

Total blood

volume

(ml/Kg) body

weight

58

78

75

70

75

Available

blood volume

(ml/Kg) body

weight

20

30

35

35

40

Clotting

time(seconds)

20

14

-

60 – 300

90

RBC Life

span (days)

45 – 68

20 – 30

-

45 – 70

-

RBC count

(cells/cumm)

7.2-9.6

7.7 – 12.5

4.5 – 7.0

4.5 – 7.0

4.5 – 6.5

Hemoglobin

(Kg/100ml)

14.8

14.8

12.4

13.6

12.5

Blood plasma

pH

7.3

7.3

7.3

7.3

7.3

Hematological Data Of Laboratory Animals

(CPCSEA guidelines)

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Annexure -I

Mice Rat Monkey Guinea pig

Rabbit

Crude protein(%min)

20.0

20.0

20.0

24.0

20.0

Ether extract(%min)

4.0

4.0

6.0

3.5

3.5

Crude fiber (%max)

4.0

4.0

4.0

12.0

12.0

Ash (%maximum)

8.0

8.0

8.0

8.0

8.0

Calcium (%minimum)

1.0

1.0

1.0

1.2

1.2

Phosphorus (%min)

0.6

0.6

0.6

0.6

0.6

Nitrogen free extract (%)

55.0

53.0

53.0

43.0

47.0

Metabolisable energy (K cal/Kg)

3600

3600

4000

3000

3000

Basic nutrients required for common laboratory Animals (CPCSEA GUIDELINES)

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Annexure -II

Diet composition for Rodents: Rat, Mice, Hamster AS PER UFAW

1. Wheat flour : 22%

2. Roaster Bengal gram flour : 60%

3. Ground nut flour : 10%

4. Skim milk powder : 5%

5. Casein : 4%

6. Refined oil : 4%

7. Salt mixture with starch : 4.8%

8. Vitamins &choline mixture with starch : 0.2%

Diet composition for monkeys, Rabbits & Guinea pigs

1 Wheat flour : 61%

2 Roaster Bengal gram flour : 28%

3 Casein : 1%

4 Refined oil : 5%

5 Salt mixture with starch : 4.8%

6 Vitamin & choline mixture with starch : 0.2%

7 Vitamin C : 50 mg/100 g diet

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Annexure –II Contd

Extra diet for some Laboratory Animals

Monkey Rabbit Guinea pig

1. Bengal gram 20g 20g 25g

2. Ground nut 15g - -

3. Plantain 1g -

4. Lucerne grass - 100g 50g

Salt mixture composition - Rats, Mice & Hamsters

Mineral Per 100 kg diet gm 1. Di calcium phosphate 1250.00

2. Calcium carbonate 555.00

3. Sodium chloride 180.00

4 . Magnesium sulphate 229.20

5. Ferrous sulphate 108.00

6. Manganese sulphate 16.04

7. Potassium Iodide 3.16

8. Zinc sulphate 2.192

9. Copper sulphate 1.908

10. Cobalt chloride 0.092

A portion 1 /2 or 1/3of the sodium chloride is mixed with the Potassium Iodide and

grind well and the mixture is stored with sufficient quantity of starch

All Minerals together : 2345.492 gm

Starch : 2454.508 gm

Total : 4800.000 gm

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1. Canadian council on Animal Care-Guide to the care and use of

experimental animals.

2. CPCSEA Guidelines.

3. Control of the Animal House environment 1976. Laboratory Animals

Handbooks.

4. Guide for the care and use of Laboratory Animals. DHEW publication

Washington 1978

5. Environmental monitoring in a laboratory Animal facility.

Lab.Ani.sci.1976; 26:592

Sources

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