cardiac+puncture

Cardiac Puncture

also located here "S:\Research\Cancer_Eradication\Discovering tumor specific antigens\Protocols\Cardiac Puncture.docx"

http://www.youtube.com/watch?v=cRx0E-bu95Q http://waweisslab.ucsf.edu/display_protocol.php?PTCID=o2uir1ozi1

Protocol from Current Protocols in Immunology http://onlinelibrary.wiley.com/doi/10.1002/0471142735.im0107s73/pdf

see also a protocol from Deborah Hansen "F:\kurt\storage\CIM Research Folder\DR\2013\5-8-13\spleen and blood collection protocols\20130507 final bleeds + spleen.xlsx"

CARDIAC PUNCTURE OF MOUSE AND RAT Materials Additional reagents and equipment for anesthesia (UNIT 1.4)
 * 1- to 3-ml syringe (mouse) or 3- to 10-ml syringe (rat), with 20- to 22-G needle
 * Serum separator BD Microtainer tube for collecting blood
 * Anesthetize the mouse or rat (most likely with avertin). Place the animal in dorsal recumbency.
 * Before inserting the needle, pull the plunger back and forth to loosen everything up.
 * Insert needle just below and slightly to the left of the xiphoid cartilage at the base of the sternum, at a 15◦ to 20◦ angle (mouse) or 20◦ to 30◦ angle (rat) (Fig. 1.7.4). In my experience, the needle often does not need to be inserted very far.
 * Advance the needle slowly, applying very slight negative pressure on the barrel of the syringe (blood will flow into the hub of the needle when the tip has entered one of the chambers of the heart). Aspirate gently until blood flow ceases. Advancing or retracting the needle tip may be necessary to obtain a maximal volume. Also slightly rotating the needle slightly seems to help greatly in some situations.
 * Remove the needle from the syringe and transfer the blood from the syringe to a BD microtainer serum separator tube (Blood Separation Tubes). Note that the blood should not be passed through a needle because this can damage some components in the blood.
 * Note that once obtaining blood from the heart becomes too difficult, a fair amount of blood from the punctured heart can be collected from the bottom of the body cavity.
 * Ensure death by cervical dislocation (UNIT 1.4) when collection is complete.
 * If it is absolutely necessary, more blood can be collected by cutting the mouse open and collecting blood from the bottom of the cavity (it may help to snip the heart)

Note: I think Andrey used a 25 guage needle one time (smaller numbers indicate larger holes)