TECHNIQUES OF BLOOD COLLECTION IN LABORATORY ANIMALS

INTRODUCTION To study the effects of test drugs. Hormones, substrates or blood cells. Terminal and non-terminal blood collection techniques. nonDeath of the animal (terminal experiment). Terminal blood collection under anesthesia. NonNon-terminal blood collection from conscious animal.

ASPECTS OF ANIMAL WELFARE

Minimizing pain & distress. Humanitarian reasons. Good scientific practice. Physiological changes associated with stress. e.g. :- increases in the blood levels of catecholamines, :prolactin & glucocorticosteriods can influence certain metabolic parameters, such as glucose, as well as the counts of erythrocytes, white cells, & packed cell volume.

TOTAL BLOOD VOLUME

Depends on species, sex, age & health ass well as nutritional conditions. Blood volume less in older, obese when compared to normal. Total circulating blood volume is in the range of 55-70 ml/kg 55body weight.

TERMINAL BLOOD COLLECTION

Exsanguination as a single process. Multiple blood sampling during terminal experiments by physical stunning or general anesthesia. Types :1) Blood withdrawal from the V.cava caudalis or the aorta after laparactomy as much blood as possible should be removed in a sterile manner. manner.

2) Exsanguination after decapitation, incision of the jugular vein or carotid artery or techniques in the slaughterhouse NonNon-sterile collection. collection.

3) Retro-orbital bleeding Retrorats

In mice, hamsters &

which can also be method of exsanguination.

NONNON-TERMINAL BLOOD COLLECTION (From conscious animal)

Not to with draw too much of blood. False results. Reduced total blood volume in animal. Reduced hemoglobin content & reduced oxygen transport capacity. Fall in blood pressure. Increased conc. of stress related hormones. Necrosis of the gastric mucosa. Individual animal should not be endangered by removal of too large volume & too frequent collection of blood. E.g. :- mice, rats or hamsters are used. :-

SINGLE BLOOD REMOVAL

Removal of 15 to 20% Accompanied by side effects such as fall in c.o or b.p. Haemorrhagic shock By the withdrawal of 30-40% 30of total blood volume. Loss of 40% causes mortality. 3-4 weeks gap between every withdrawal. Hypovolaemic shock symptoms fast pulse, pale mucous membranes, hyperventilation, normal body temp including cold skin & extremities.

MULTIPLE BLOOD REMOVAL

Not exceeding 1% of total blood volume every 24 hr (0.6 ml/kg/d). Symptoms of anemia pale mucous memb of conjunctiva or inside the mouth, intolerance to exercise & an increased respiratory rate. Anemia can be easily detected by PCV, ECR, Hg. Anemia treated with iron & vitamin B12.

TECHNICAL ASPECTS OF BLOOD REMOVAL

0.1 ml Tip of the tail. Fresh capillary blood. E.g.:E.g.:- Blood glucose or total radioactivity after the administration of radio labeled drugs. Guinea pig cardiac puncture under general anesthesia. In large animals blood collection by superficial vein. Locate the vessel accurately before insertion of needle or catheter.

Bore of the needle should be as large as possible

to ensure rapid blood withdrawal. Continuous pressure should be applied immediately and maintained for 30 sec. Monitored 15mins later.

PERMANENT VENOUS CANNULATION

By chronic cannulation. Particularly in rats. A few days after the implantation of catheters, hormone levels are normal. Housed alone. Tethering restricts normal movements such as lying on the back and rolling over. At the time of experiments longer catheter is attached.

SHORT -TERM CANNULATION

A butterfly needle.

Clotting can be prevented by repeatedly filling the catheter with saline containing heparin.

RETRORETRO-ORBITAL BLEEDING
Orbital puncture. Orbital venous plexus. Tail less animals e.g. :- hamsters also rats & mice. :Pasteur pipettes, micropipettes or micro capillary tubes. Rotating movements through the conjunctiva laterally, dorsally or medially of the eye to the back wall of the orbit. Retrorbital haematoma with subsequent pressure on the eye cannot be excluded

CARDIAC PUNCTURE

Performed in guinea pig, gerbils & hamsters. Difficult to collect blood by alternative methods except retroretro-orbital bleeding. Performed under general anesthesia.