It took me half an hour to separate serum from Guinea pig blood by allowing blood to stand still in a slant position at room temperature followed by centrifugation. I need a quicker means of serum separation to produce high-titre complement.
For separating serum from the blood cells you should use a tube without any anticoagulant , sterile empty tube will be fine , after taking your sample , say 5 ml or so leave the tube in a standing position for about 20-30 minutes ( it can take shorter time than this so you can check it preiodically ) after that you will find your blood to be clotted , centrifuge at 20 c degree , 1500g for 10 minutes then remove your serum very quickly and flash freeze it in -80 c to preserve your complememnt for next assay.
Hello,
Agree with Olubayode. Also, note that you should allow blood to clot at room temperature, or, as I remember, 37 degree centigrade for human samples. Low temperature will lower the speed of fibrin formation and blood clotting.
the best method ever that prescribed by Alton et al, 1988. which can be summarized as follow:
1- bleed at least 4 adult guinea pigs, preferably many more
2-collect the serum as soon as it has separated from the clot of incubated blood at 37 degree centigrade for 15 min, separation should not take more than 1 hour (as complement is the most fragile part in CFT method)
3-pool the sera and centrifuge at 900 xg to remove any erythrocyte. the clear serum produced is then used as complement
4-when the adequate facilities for freezing or drying complement are not available, it may be preserved by richardson's method and when preserved will maintain its titre for about 6 month if stored at 0-4 degree centigrade. even at room temperature, loss of titre is not rapid
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