Hi Rolayo

Because the volume of blood is small, instead of trying to separate buffy coat by centrifugation, you may want to use a membrane filter with 3 micron diameter pores. Red cell will go through them under negative pressure but the white cells will be retained on the surface of the filter. You can then rinse the filter with saline to retrieve the white cells. I did a similar experiment to visualise white cell fragments in filtered blood. Very effective method to capture small number of white cells and fragments. You may want to have a look at this publication-

A new method to study the efficiency of third generation blood filters - PubMed (nih.gov)

Br J Haematol.,1993 May;84(1):175-7. doi: 10.1111/j.1365-2141.1993.tb03045.x.

Good luck

Siva

Is it not possible to use microhematocrit ?

Thank you @Muttuswamy. My target is Trypanosomes not the white cells.

@Ramesh, I need larger quantity of buffy coat beyond haematocit volume. Thank you for your suggestions.

I am not a parasitologist, pardon me for my ignorance and tell me how about the utility of an old technique given in a recent artical

Hello Sir,

Would recommend diluting the 0.5 ul of whole blood with an equal volume of PBS and then overlay this mixture onto 1.0 ml of Ficol-Hipaque solution.

Then spin this mixture for 20 minutes at 1800 rpm, keeping acceleration rate: 4 and De-acceleration rate: 4. This protocol I had standardized for patient samples where the sample volume was very less.

Regards.

Thank you @Priya