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I did try a few dilutions of the primary antibody (1/400, 1/200, and 1/100) with no beautiful membranous staining. We are really looking for something very distinct in order to show colocalization with our experiments' target protein. I've attached an image of the 1/100 dilution at a 20x1.5x magnification that shows maybe some membrane staining, but a lot of background. Any suggestions?
Also, I forgot to include my promotional code in my abreview for one free antibody: xxx
Can you please let me know what I need to do in order to get my next antibody free?
Asked on Nov 01 2012
Thank you for your reply.
As for the staining, the image you sent seems a bit dark. Also, the Abreview image seemed a bit dark - independent of which color you had used. I'd suggest checking the secondary to ensure it is bright enough or to avoid long over exposure in the microscope.
You might also increase the blocking time to reduce the background, and switch to BSA or milk.
Using a positive control (human sample) - if available - could help with optimizing the antibody as well.
Another option to try could be a different fixation method (methanol or acetone).
I hope this is of help.
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Answered on Nov 01 2012