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Author Topic: blocking peptide  (Read 3113 times)

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Offline yuter

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blocking peptide
« on: May 06, 2003, 07:49:16 AM »
In my IHC for substace P on nasal mucosa,the brown color (DAB) of the slides with blocking peptide is even darker than those without blocking peptide(Santa Cruz).So I am confused why does this happen(is it because of the high affinity of the blocking peptide to the tissue)?
I ask a pathology professor and she answer me firmly I must have something wrong in my procedure,it's impossible.But another antibody for IHC under the exact same procedure and condition (I performed them simultaneously) are normal.Any one have ever meet the problem?Thank you!

blocking peptide
« on: May 06, 2003, 07:49:16 AM »

davidtest

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blocking peptide
« Reply #1 on: May 06, 2003, 10:50:32 AM »
If you can post the detailed procedure you are using for the absorption control, it will be very helpful for figuring out the points of problems.

David

Offline ihcwor2

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blocking peptide
« Reply #2 on: May 09, 2003, 09:25:37 AM »
Possible reasons:

1) The concentration of primary antibody in blocking peptide mixture is higer than the concentration of primary antibody without blocking peptide.

2) The ratio of primary antibody and blocking peptide in the mixture is incorrect (normally it should be around 1:5).

3) The mixture should be incubate at room temperature for at least 1 hour before applying to sections.

4) You should wash sections throughoutly after incubation of the mixture.

Hope this helps.

Richard

blocking peptide
« Reply #2 on: May 09, 2003, 09:25:37 AM »