Immunohistochemistry free floating abreview for Anti-GAD65 + GAD67 antibody

Immunohistochemistry free floating

Rat Tissue sections (Brain)

Brain

24 hour(s) and 0 minute(s) · Temperature: 4°C · Diluent: 5% normal goat serum in 1x PBS

1/2000

1/250

Non-Abcam antibody was used: Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed S
Host species: Goat
Clonality: Polyclonal
Conjugation: Alexa Fluor® 647

My tissue was fixed with 4% paraformaldehyde, then a 30% sucrose solution mixed with PBS and 0.1M sodium azide before being flash frozen in methyl butane, stored at -80, sliced, and then underwent IHC and free floating sections. My goal was to see clearly labeled cell bodies with minimal background for a cell body colocalization study. In the cortex, some cell bodies were visible but it was incredibly difficult to pick them out because of the high background. I tried a 5% blocking solution for 30 mins or 60 mins, and mixed the primary antibody in a 5% blocking solution with no luck. I tried dilutions of 1:500 (not pictured because I can't upload more than 1 image), 1:1000 (not pictured), 1:2000, 1:4000 (not pictured), and 1:8000 (not pictured) with no clear improvement.

Thank you for participating in our AbTrial program and your valuable feedback. As the product had not yet been tested in this application we were unsure what the result might be and we appreciate the time and effort that the reviewer has put in to testing this product in this application.