facs analysis - (Jun/18/2011 )
i am doing FACS analysis for quite some time but not getting good result as expected by my boss.
i am checking the expression level of my receptor protein after bacterial infection compared to no infection .
i am getting very slight inc in expression .
i am attaching a file for that .
but my boss is expecting a more % of shift.
is my data valid or not .
attaching some file
I think based on the result on the attached file, there is no significant difference. but
why your boss thinks it should change more %? other experiment indicate the difference?
One thing that I can think of is, if the cell is adherent cell and you are using trypsin before FACS,
Trypsin might be affecting the receptor.
If you have positive control (or condition which increase the receptor expression) you can test that.
Otherwise I will test expression level after different time points after infection.
i don't use trypsin .
i just scrap the cells using scraper.
how to troubleshoot it i have no idea and my boss is always after me for the same
Have you ever this experiment with different methods such as immunocytochem??