# Problem in IHC PROTOCOL

**URL:** <https://scientistsolutions.discourse.group/t/problem-in-ihc-protocol/759>\
**Category:** Immunohistochemistry\
**Created:** [January 16, 2025, 9:02am UTC](https://scientistsolutions.discourse.group/t/problem-in-ihc-protocol/759 "2025-01-16T09:02:21Z")\
**Posts on this page:** 3\
**Page:** 1

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**Author:** ![BarryArmstrong](https://avatars.discourse-cdn.com/v4/letter/b/8e7dd6/32.png) [@BarryArmstrong](https://scientistsolutions.discourse.group/u/BarryArmstrong)\
**Post date:** [January 16, 2025, 9:02am UTC](https://scientistsolutions.discourse.group/t/problem-in-ihc-protocol/759/1 "2025-01-16T09:02:21Z")

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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

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**Author:** ![excalpath](https://avatars.discourse-cdn.com/v4/letter/e/c67d28/32.png) [@excalpath](https://scientistsolutions.discourse.group/u/excalpath)\
**Post date:** [March 13, 2025, 2:55pm UTC](https://scientistsolutions.discourse.group/t/problem-in-ihc-protocol/759/2 "2025-03-13T14:55:44Z")

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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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**Author:** ![Pundir-nbrc](https://yyz2.discourse-cdn.com/flex004/user_avatar/scientistsolutions.discourse.group/pundir-nbrc/32/131_2.png) [@Pundir-nbrc](https://scientistsolutions.discourse.group/u/Pundir-nbrc)\
**Post date:** [August 18, 2025, 1:06pm UTC](https://scientistsolutions.discourse.group/t/problem-in-ihc-protocol/759/3 "2025-08-18T13:06:46Z")

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What is your host for the secondary antibody? Use the serum of the secondary antibody host species along with BSA for blocking non-specific signals.