The different results obtained were in different days, using different samples of blood? I didn't understand... 0.1% is the concentration of erythrocites in the test or the concentration of the essential oil?

Below some hypothesis that could justificate these results:

- Did you standardize the amount RBCs in the test?

- In each assay that you repeated, did you perform in replicate?

- The controls (water or Triton X-100, and saline or PBS) worked well? They produced the same absorbance every time you repeated the test?

- Did you use the same sample for the testing in all repetition? If yes, could the time influenciate in the stability of oil? If no, maybe the concentration is very low so its difficult to weight or pippete the same amount every time.

It's a strange situation. If you could provide some more details, maybe we could resolve this problem.

Best regards,

MSc. Juliana Felix.

Thanks Juliana for your response.

0.1% is the concentration of the oil. The amount of RBCs in the test was 20 ul.  I did repeat the assay on separate occasions using the same sample. The positive and negative control work well and gave me the same results each time (I used PBS for negative and water for positive) and I incubated the sample for 2 h at 37 c.  

Thanks 

Dear Fahd Sam Ali:

ensures the same concentration of cells to the volume of 20 ul

pipetting should be soft, the RBC can be lysed by pressure

good luck

Hi Fahd. I just saw your question and the comments, so this one is a bit delayed. Besides the notes already made, I'd suspect that measuring 20 ul blood may have different number of red blood cells. It would help to control this as a variable to count the cells, so you add the same number of cells each time. Either human or animal origin, blood may come a bit different from day to day.

HI,

Which is the commonly used blood group for hemolysis assay? If you prefer a particular group, why is it?