Blood Collection - Rats

Revision Date: June 8, 2022
Responsible Office: Animal Research

1. Acceptable Collection Volumes
2. Procedure Preparation
3. Procedures
4. Post-procedure Monitoring and Care

Blood Collection from Rats

Any of the individual procedures described below, when performed properly by trained individuals do not cause more than momentary/transient pain or distress, with the exception of retro-orbital blood collection (which is performed under general anesthesia).

1. Acceptable Collection Volumes

A maximum of 10% of total blood volume may be collected weekly. If the maximum volume is collected, to 1.0-2.0ml of sterile saline may be administered subcutaneously to rehydrate the rat.

2. Procedure Preparation

a. The work space will be clean and organized with all blood collection equipment to minimize the manipulation of the rats.

b. If anesthesia or sedation is used, it will be a regimen approved in the Anesthesia Policy, Guidelines and Procedures.

3. Procedures

Retro-orbital

a. Rats must be anesthetized to alleviate potential pain or distress associated with the procedure.
b. If serial collections are necessary, they will be taken from alternating eyes at an interval of no less than 7 days with no more than 3 procedures per eye (during the lifetime of the animal). Other methods will be used if blood must be collected at a greater frequency.
c. A topical ophthalmic anesthetic, e.g. proparacaine or tetracaine drops may be used.
d. The animal will be immobile, exhibit no response to a firm toe pinch, and respiratory rate will be slightly reduced.
e. The rat is restrained with the facial skin pulled taut while grasping the skin at the back of the neck (the scruff) to restrain, resulting in a slight protrusion of the eyes. Care is taken to not compress the thoracic cavity.
f. The tip of the pipette (micropipettes, capillary tubes, or similar blood collection device) is gently inserted at the medial or lateral canthus, at an approximate 45° angle into the space between the globe and the lower eyelid.
g. With a gentle thrust and rotation motion past the globe of the eye the tube will enter the slightly resistant retro-orbital plexus. The globe itself remains uninjured. As soon as the sinus is punctured, blood enters the tubing by capillary action. Additional pipettes may be used to collect the necessary volume.
h. When the desired amount is collected, gentle pressure with clean gauze is placed on the eye to ensure hemostasis.
i. If an eye is damaged, this eye will not be used again for blood collection. VCS should also be notified.

Saphenous Vein

a. Light sedation may be used, based on comfort level of personnel.
b. Awake rats will be placed in an appropriately ventilated restraining tube with its head covered and hind legs free. If anesthetized, the respiratory rate of the rat is monitored carefully for depth of anesthesia.
c. Fur may be removed from the lateral surface of the lower leg with clippers or carefully with a scalpel blade.
d. The area may be wiped with 70% alcohol.
e. Petroleum jelly, eye lubricant, or a comparable product may be applied to the puncture site to prevent migration of blood into the surrounding fur.
f. To dilate the vein, the hind limb is gently extended, and pressure applied to the thigh area proximal to the puncture site.
g. The vein is punctured with 23G sterile hypodermic needle.
h. Drops of blood are collected, as they appear, in micropipettes or small blood collection tubes.
i. Frequent serial collection may be achieved by moving proximally, up the saphenous vein and using both hind limbs.

Tail Vein Nick

a. Light sedation may be used, based on comfort level of personnel.
b. The rat is placed in a restrainer and the tail stabilized using the thumb and forefinger of the non-dominant hand.
c. The end of the tail is gently rotated to expose the lateral tail vein on either side.
d. The area is wiped with 70% alcohol.
e. Approximately 2-3 cm from the tip of the tail, a small nick is made over the lateral tail vein using a sterile scalpel blade or 21G-23G hypodermic needle. When the nick is made deep enough, blood should start welling up from the nick immediately.
f. Drops of blood are collected, as they appear, in micropipettes or small blood collection tubes.
g. Frequent serial collection may be achieved by moving proximally, up the tail vein and by using the vein on the opposite side of the tail. The original site may be used again after allowing 7 days for healing.

Tail Snip

a. Light sedation may be used, based on comfort level of personnel.
b. The rat is placed in a restrainer or handheld with an assistant to restrain the rat.
c. The area may be wiped with 70% alcohol.
d. Up to 2mm of the tail tip is excised.
e. Drops of blood are collected, as they appear, in micropipettes or small blood collection tubes.
f. Frequent serial collection (minutes to hours) may be achieved by removing the “scab”.

4. Post-procedure Monitoring and Care

a. For all methods of collection, gentle but firm pressure is applied to the collection site until hemostasis has been achieved.
b. Rats that have been anesthetized using ketamine/xylazine are placed in a bedding free cage or a cage with a paper towel on top of bedding with thermal support provided and monitored until animal is fully recovered. Refer to Thermal Support of Anesthetized Laboratory Mammals and Birds Policy for more information.
c. Significant damage to the collection site will restrict use of the original site. That site will no longer be used, and additional blood collections will be obtained via a different route unless in a terminal procedure. VCS will be notified, and a treatment plan implemented.