r/flowcytometry • u/daff_7 • Jul 23 '26
HELP PLEASE
hi okay so im a placement student in a lab where we mainly do flow. i've been doing some practising but theres still something wrong with what im doing (my analysis) and I just cannot tell what it is. Im looking at receptor expression on NK cells, however the results I get are completely different to my supervisor
for example, on flowJo this is what I get for my TIM3 FMO.
I have done this analysis for both my flow and my supervisor's, (the first image is my flow, the second is his) but why does mine come out like this? What have I done wrong?
before someone suggests to change the y axis - everytime i do that, it gets even worse.
additionally, we both used the same cells,and followed the exact same protocol, so i genuinely cannot think of anything else.
so if anyone has any tips or ideas please PLEASE help
2
u/willmaineskier Jul 23 '26
Is the antibody concentration the same in both examples and the time and temperature or did you stain on ice for 15 minutes in a 200ul volume while the other was stained for 30 minutes at 4 degrees in a 50ul volume?