Problem in IHC PROTOCOL

Cytology, Histology & IHC Immunohistochemistry
1

Hi all

I am doing IHC on rat heart sections using goat primary antibody for collagen 1, 3 & TGF beta with secondary antibody antigoat HRP conjugated and DAB as a substrate. I am using the primary and secondary Ab at the conc. of 1:200. I m not gettin any kind of signals although my cunterstaining is good and there is no background stain. I have few querries:

Since I m using Abs generated in goat can i use 5 % normal goat serum for blocking. Currently I m using 3% BSA as a blockinf agent.

Ours vecta stain kit is for mouse and rabbit secondary Abs. Can I use them with my Abs which are generated in goat.

If cannot then what can be the alternative procedure if any

2

Since your antibody is hosted in goat you will need to use a goat detection kit. Ones for rabbit and mice will not work. Always use detection for the host animal of your antibody.

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