I have been taking blood samples from Wistar Rats and I had some problems with plasma collecting. After centrifugation, plasma still showing a pale red. I am using a decapitation prodecure in order to get the blood, and some colleagues have pointed it out as the root of the later disturbed plasma. Nevertheless, I actually think that my EDTA solution is the problem. I am using the proportion 0.5ml EDTA disodic dihidrate (1%) into distilled H20 for 5ml blood. I had no problems with the coagulation, but after centrifugate (2000rpm, 15min, 4ºC) my plasma sample still being red (palid red). I want to know if this EDTA % and ratio EDTA/Blood is correct, or if I shoud use another one (For example, EDTA 0.5M, or 6%). Thus, I would like some expert in such issue explain me his/her ENTIRE protocol (EDTA type, solvent, PH correction, EDTA temperature at the moment of blood collection, blood extraction procedure, EDTA/ Blood ratio, rpm and time of centrifugation, etc). Thanks.
Hello,
I think the problem is the water you use. The isotonic solution causes hemolysis which explains that color. I recommend an isotonic NaCl solution to dilute your EDTA. In the tubes for human biology concentration EDTA K2 is 1.8 mg / ml. Centrifugation 10 min at 1700 G 20°C.
Cordially
I agree with Valery, you need to use isotonic buffer (for example - PBS) for manipulation with RBCs.
- Badges
- Science method
- Similar topics
- Chemistry
- Biochemistry
- Biochemical Methods
- Protein Purification Techniques