Preparing Serum from Whole Blood

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by Michael Koelle, from Harlow and Lane "Antibodies: A Laboratory Manual"

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1. Veterinary services people often will do bleeds for you and give you the fresh blood in glass vacutainer tubes, leaving you to prepare serum from it.

2. Put the blood at 37° for 30-60 min to clot.

3. Pull the rubber stopper off the top, and use a long Pasteur pipette to "ring" the clot, i.e. separate it from the sides of the glass tube. Alternatively, just invert the tube; if the clot pulls free from the glass walls of the tube, then you don't have to ring the clot. Set the tubes at 4° overnight to allow the clot to contract.

4. Decant the serum away from the clot into a new centrifuge tube. Some red blood cells and bits of clot will come along with the serum. Spin the serum 3,000 RPM for 10 min to remove the remaining clots/red blood cells and other insoluble material.

5. Make nicely labelled aliquots and store at -80°. The antibody should be stable for many years. A working solution can be stored at 4°. If azide won't interfere with your applications, you should add 5-10 mM sodium azide to this working solution tookeep bacteria from growing in it.

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