Blood Collection: The peri-orbital sinus of the mouse may be used as a source of venous blood.  The mouse has a large peri-orbital venous sinus that fills the bony orbit of the eye.  Blood can be collected from either medial or lateral canthus approach.

The Duke Institutional Animal Care & Use Committee has determined that peri-orbital bleeding of anaesthetized mice is an acceptable procedure in the hands of a skilled technician.  Personnel MUST NOT attempt this technique until they have had opportunity to learn from experienced personnel.  This procedure will require specific justification over why a less invasive procedure cannot be performed.

• Equipment:

  • Paper towels (or similar)

  • Kimwipe, kleenez (or similar)

  • Microhematocrit tubes, 200ul pipettes ot tube of similar diameter.

• Procedure:

  • Lateral Canthus:  Pick-up the mouse and restrain it in one hand.  Inset the tube into the lateral canthus.  The tube should be at about a 30 degree angle to the side of the head.

  • Medial Canthus:  Place the mouse on a table or cage lid in lateral recumbancy.  The body of the mouse is restrained against the table with the palm of the hand.  The thumb and forefingers of the same hand restrain the hand and gently open the eyelids to expose the eye.  Insert the tube into medial canthus and hold it at a 30 degree angle to the nose.

  • Both Procedures:  A microhematocrit tube or pipette is inserted through the conjunctiva and into the orbital sinus by quickly rotating the tube.  The eye is not damaged because the tube passes under the eye.

    • Blood flow may be increased by changing the angle of the tube slightly.

    • After the required amount of blood is obtained, the tube is withdrawn and bleeding usually ceases by the eye (orbit) pressure alone.  If necessary, hemorrhage can be controlled by direct pressure with a Kimwipe or Kleenex over the eyelids.

     

• Comments:

  • Small amounts of blood (30-80 micro liters) can be obtained from the orbital sinus of mice as young as 14 -16 days of age.

  • Larger amounts of blood (up to 0.5 ml) can be obtained from orbital sinuses of older mice if tubes containing anticoagulant are used.

  • No more than 10% of the animals blood volume should be collected every four weeks (for a normal adult mouse this means not more than 250 ul over a four week period).  Any large quantity of blood collection should be accompanied by subcutaneous fluid (warmed saline) replacement of the same volume.

  • Orbital bleeding can be repeated within hours if the amount of blood removed at any time is relatively small.

  • Orbital bleeding of mice less than 14 days is not considered a routine procedure, but can be safely performed