Comments & suggestions  should be directed to jroese@lssu.edu Visit my website Blood Collection A safe maximum for a single sample is 7 ml/kg body weight (.07 ml or 70 Fl per 10 grams of body weight) taken every 2 weeks.  Animals on chronic studies requiring multiple blood samples may need to have their hematocrit monitored.  Several techniques may be employed to collect blood samples from mice. Retroorbital bleeding is one of the most commonly used techniques for routine blood collection. The mouse must be anesthetized for this procedure.  Pressure is placed on the top and bottom lids of one eye to keep the eye open and pushed forward slightly.  A glass microcapillary tube is placed in the medial canthus of the eye at a 30°- 45° angle toward the back of the eye.  Use firm, steady forward pressure and rotate the tube between the thumb and forefinger to cut through the conjunctiva at the back of the eye and enter the retroorbital sinus, at which time blood should flow into the tube.  If no blood is obtained gently back off on the position of the tube.  After collecting the sample, close the eyelids and apply pressure with a piece of gauze until bleeding stops (hemostasis).  A small amount of ophthalmic ointment containing an antibiotic may be placed on the eye after bleeding has stopped to act as a "bandage" and help prevent infection.  If the technique of the sampler is good, the eye should not be damaged.  If frequent samples must be collected, alternate the eye used for sampling. The tail vein is another site commonly used to collect small blood samples from mice.   Older mice should be anesthetized for the procedure, even though blood is harder to obtain from the tail vein when blood pressure drops under anesthesia.  A small nick is made in the tail vein with a scalpel blade.  Care must be taken to avoid cutting into the artery or amputating the tail.  Blood can be collected using a microcapillary tube or may be allowed to drip into a small eppendorf tube.  After blood has been collected the tail incision should be compressed with a piece of gauze until hemostasis occurs.  If bleeding is difficult to stop a silver nitrate stick may be applied.  This technique can result in scarring to the tail. The saphenous vein is rapidly becoming the technique of choice for blood collection for many investigators. This procedure does not require that the mouse be anesthetized to collect a blood sample, and is much less invasive than the two previously mentioned techniques.  A tube can be used to restrain the mouse, and the hind leg is extended by applying gentle downward pressure just above the knee.  The hair over the tarsal area is shaved with clippers followed by a number 11 scalpel blade, and the saphenous vein is pricked with a needle (25 gauge is usually adequate). Blood can be collected in a microcapillary tube.  Smearing a small amount of silicone grease over the area to be punctured helps to prevent the blood from coming into contact with the fur and minimizes blood clotting.  When blood has been collected, gentle pressure applied with a piece of gauze should be used to effect hemostasis. Terminal bleeding procedures are commonly used to collect larger volume samples from anesthetized mice at the time of euthanasia.  In one technique, the brachial vessels are exposed by removing the skin in the axilla and the vessels are then cut.  Pooling blood is collected using a pipette, and blood collection must be done quickly as clotting is initiated by contact of blood with  tissue.  Terminal bleeding can also be done by collecting blood directly from the heart.  Heart puncture may be done “blindly” by directing a needle into the thoracic cavity from the outside after palpating the beating heart at the level of the 5th  intercostal space.  Alternatively, the chest cavity may be opened so that the heart can be visualized, at which time the needle can be introduced into the right ventricle to collect blood. Blood Collection from the Tail Vein. Warm the tail in a water bath and make a small nick in the tail with a sharp scalpel blade.  Collect the bood with a microcapillary tube. Step 1. Place the animal in a restraining device and gently extend the thigh.