Hi Arianna,

I confirm you, Hek293a should not express BDNF. So two possibilites:

- You have contaminations during your experiment, ELISA is very sensitive, Any supplementation in your medium which could contamined? You can also do a negative control with only PBS or H2O. You could test another detection technique if you can.

- Your cell line has derived or is not exactly Hek293a. How do you get it? by another lab? use in your lab since more 10 years? Ordered to a company?

Hope these insights will help you,

Regards

Nicolas

Many thanks for the reply, Nicolas!

I just got this cell line from another lab in the campus but I am pretty sure they have been working with them for a while.

I will try to run also some negative controls (blank) next time, but I am quite sure that there is something in the medium which is contaminating the results because the standard curve was fine...btw, I will figure it out!

Cheers,

Arianna

Hi Arianna,

Yes it could be that, and if it is that, the question is if I understand well your experience, why you detect a higher signal with medium + non transduced cells than with medium alone?

If you get the cells from another lab, I suggest you to ask them the history of the cell line, I speak from experience; during my thesis I get a cell line from another lab which has been not what it was write on the batch and which realy looks like to the expecting cell line.... three months to realize that.....

Looking forward to hearing from you about your next results.

regards

Nicolas

Hi Nicolas,

Yes, that's right. So I was thinking, in your opinion is possible that I took some cells or cells debris while I was aspirating the medium off?

I haven't centrifuged the supernatants, I just got them and stored -80C.

Should I have centrifuged all my samples before freeze them? Should I centrifuge my samples next time?

Also, thanks for your help and advice. I will do double check with the cell line.

Cheers,

Arianna

Hi Arianna,

Yes if you do not centrifugation, you aspirate some cells and debris, so it is better to do centrifugation if you want to be sure to test only the medium.

Please let me know your next results.

Cheers

Nicolas