CLEANING AND TAGGING OF LABORATORY ANIMALS

Cleaning

• Change rabbit & rodent bedding. 
• Change & sanitize cage weekly or up to twice weekly for rats and spot changed later in the week if needed. 
• Change pans are sanitize twice weekly or biweekly. 
• Sanitize all water bottles, stoppers, and sipper tubes weekly, replace bottles/replenish feed as needed. 
• Sweep if necessary. 

Tagging (Identification)

Small rodents

• Rodents must be identified to the cage level, by protocol number, investigator, species, strain, sex, contact name and number, date of arrival or birth and age. 
• Rodents may be identified individually by tattooing, microchips, ear punches, ear tags, clipping hair, dyes, magic marker. 

Rabbit & guinea pigs

• Guinea pigs, chinchillas and rabbits must be identified to the cage level similar to small rodents. 
• Individual animals may be identified with tattoos, microchips, dyes, hair clipping or Magic Marker.

FEEDING OF LABORATORY ANIMALS - SMALL RODENTS (MICE,RATS AND HAMSTER)

Feeding of laboratory animals - Small rodents (Mice, rats, hamster)

• Feed rodent chow.
• Feed a high fat chow for breeding animals for extra energy, low fat diet for non breeding animals
• Put food in the mouse food hoppers at least 1/2-2/3 full and fill completely weekly. 
• Never recycle food.
• Give mice and gerbils 3-6 grams of food per day- approximately one biscuit per animal per day; hamsters 15-20 g, or 3-4 biscuits/animal/day; rats 20-50 g or 4-10 biscuits/animal/day. 
• Pellets provide something to gnaw on and help wear down the continuously growing incisor of rodents. Powdered diets usually require addition of a gnawing substrate or monitoring of tooth growth. 
• Place food pellets on the bottom of the cage for hamsters with litters.
• Some diets need to be sterilized. If this is done by autoclaving, it is essential to use an autoclaveable chow that is formulated to be nutritionally adequate after being exposed to high temperatures. Irradiated diets are already sterile and do not require autoclaving. 

Watering - Small rodents 

• Re-fill bottle that are less than 1/2 full.
• When filling, leave 1 to 1-1/2" of air space in the top of the bottle so it functions normally. 
• Replace if the water bottles leaks.
• An adult mouse or gerbil consumes less than 10 ml water per day; hamsters 25 ml/animal/day; rats 20-75 ml/animal/day. A water bottle holds 500 ml. 
• Use potable water to fill bottles. 
• Mice less than 6 weeks of age or 20 g, unweaned animals, and animals within the first two weeks of weaning must have a free-flow rather than a ball-bearing sipper tube in their cage water bottle, unless otherwise specified.
• Sanitize water bottles and sipper tubes weekly. Soak sipper tubes in a in a detergent-disinfectant for an hour to remove debris. Check the function of the ball bearing in ball-bearing sipper tubes. If the bearing sticks, discard the tube. 

Feeding - Rabbits & guinea pig

• Check food hoppers every day for caked or dusty feed. Discard caked feed and "fines" as necessary. 
• Check feed labels as guinea pig & rabbit food all look similar. 
• Rabbit feed with high fiber may be used to prevent obesity and GI obstructions. 
• Guinea pig feed has Vitamin C added. 
• Guinea pigs eat up to 35-60 g daily, rabbits will eat 100-250 g/day, but should be given no more than 120 g (4 oz or 1/2 cup) per day to prevent obesity. 
• Excessive feeding of produce for enrichment may cause diarrhea. 

Water - Rabbits & guinea pig 

• Sanitize all water bottles, stoppers, and sipper tubes weekly. 
• Rabbits will drink up to 500 ml water daily, guinea pigs as much as 50-100 ml daily. 
• Put at least 2 water bottles for rabbits and guinea pigs. 

Guinea pigs are messy drinkers and cages can get wet very quickly

HANDLING OF LABORATORY ANIMALS - MOUSE,RAT,HAMSTER,GUINEA PIG, AND RABBIT

Mouse

• Wear gloves to prevent allergies. 
• Catch and lift mice by the tail. 
• Grasp the tail with the thumb and forefinger or by the used of smooth-tipped forceps. 
• Place a second hand under their feet (for pregnant mice). 
• Hold mouse by the tail and placed on a table or other surface and grasp the loose skin over neck and shoulders with thumb and fingers for a more control grasp.
• Hold the tail or the tail and rear leg by the third and little fingers of the same hand or with the other hand to improve restraining.
• Put back mice in cage by lowering it into the cage and release upon contact with bedding.
• For mice less than two weeks of age, grasp by the loose skin over the neck and shoulder with thumb and forefinger or smooth tipped forceps.
• Avoid handling neonatal mice.

Rat

• Wear latex gloves to prevent the development of allergies due to direct contact with animal allergens. 
• Place the hand into the cage to allow exploration by the animal prior to touching. 
• Initial gentle stroking of the animal followed by gradual grasping the animal will prevent startling the animal and initiating an aggressive response. 
• Avoid approaching the animal from the front.
• Grasp the whole body with the pal over the back, with forefinger behind the head and the thumb and second finger under opposite axilla. 
• Holding with one hand is usually adequate for control, but the tail, rear legs or lower part of body may be held by the other hand for close control, treatment or examination 
• Use both hands for rats weighing over 350g. 
• Young rats may be handled like mice when body size does not permit ease of handling within the hand. 
• Avoid lifting by the tail as they may strip the skin from the tail. However, the "base" of the tail may be grasped with the thumb and forefinger.

Hamster

Method I

• Form a cup with your hands and place it over the hamster.
• Gently press your palms against the animal as you pick it up.

Method II

• Grasp the loose fold of skin behind the neck with your thumb and index finger. Cup your other hand under the animal’s rump & grasp the hind legs between your thumb and index finger.
• Carry the hamster in the same position in which you picked him up.
• For injecting the animal in the mid section, merely stretch it out.

Caution: Always handle hamster over a bench or table because they are liable to jump from your grasp and injure themselves.

Guinea pig

• Wear latex gloves to prevent the development of allergies. 
• One hand is gently placed dorsally over the thorax or ventrally under the thorax and the other hand should be used to support the animals hindquarters. 
• Do not to apply to much pressure over the thorax to avoid damaging the viscera or compressing the lungs thereby compromising respiration. 
• After grasping the Guinea pig secure it by wrapping in a towel or holding against your body will lessen the frequency of struggle. 
• Do not attempt restraint by solely grasping the skin. The lack of loose skin in Guinea pigs will result in hair depilation if this technique is utilized. 

Rabbit

• Wear latex gloves
• Held the rabbit in a way that directs their hind feet away from the handler's body.
• Grasp the loose skin over the neck and shoulder with the head directed away from the holder is the best method of initial restraint. 
• Support the lower part of the body by the the other hand to prevent serious injury to the rabbit's
• Never restrained or lifted by the ears.
• Immediately place the rabbit on a solid surface if it begin to struggle violently.
• Complete restraint should be accomplished before attempting any procedures.

LAB - PACKED CELL VOLUME TEST (PCV)

Introduction:

Haematocrit is also known as Packed Cell Volume (PCV). The packed cell volume (PCV) is the measure of the ratio of the volume occupied by the red cells to the volume of whole blood in a sample of capillary, venous, or arterial blood. The word hematocrit means “to separate blood”, which underscores the mechanism of the test because the plasmas and blood cells are separated by centrifugation.

Objective:

To measure the ratio of the volume occupied by red blood cells to the volume of whole blood in a sample of venous blood.

Equipments:

1. Microhaematocrit tube
2. Micorhaematocrit centrifuge
3. EDTA venous blood
4. Plastercene
5. Microhaematocrit reader

Procedure:

1. The blood is mixed well.
2. The blood is filled into the haematocrit tube until it is 4/5 of the blood.
3. The other end of the capillary tube is sealed with plastercene.
4. The capillary tube is placed into the microhaematocrit centrifuge.
5. The capillary tube is centrifuged for 5 minutes for 12000 rpm.
6. Read and record the results by using microhaematocrit reader.

Questions:

1. Define haematocrit.
2. What is the other term for haematocrit?
3. Name three layers of centrifuged blood.
4. Why it is important to read the microhaematocrit soon after the centrifuge stops?

LAB - WHITE BLOOD CELL (LEUCOCYTE) COUNTING

Introduction:

A white blood cell count estimates the total number of white cells. It is important in the diagnosis of disease especially when accompanied by a differential white cell count.

Two method to evaluate total number of leucocytes

1. Manual method
2. Automation technique

Reagents:

Dilution solution:

Tuerk dilution solution: 2% glacial acetic acid

®       Glacial acetic acid                  

®       Distilled water

®       Gentian violet

Principle:

Acetic acid will lyse RBC, so WBC only can be count.

Procedure:

1. Using a red blood cell Thoma pipette draw well mixed blood and fill till the 0.5 mark. Clean the tip of the tube.
2. Now draw WBC diluting fluid to the 11 mark which is 1:20 dilution.
3. Mix well for 3 minutes.
4. Discard the first three to four drops of mixture and gently fill in the mixture into the Neubauer chamber.
5. Under 10X magnification, scan to ensure even distribution.
6. Count the red blood cells using 40X objective in the 80 smallest squares as indicated in the diagram of the chamber.

Questions:

1. What are the factors that may occur for the wrong counting of white blood cells?
2. Define leucocytosis.
3. What is the normal white blood cell range for adults and babies?