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Author Topic: Citrate buffer antigen unmasking  (Read 9576 times)

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Offline Ghuman

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Citrate buffer antigen unmasking
« on: October 19, 2003, 03:55:57 AM »
Hi

I have used citrate buffer (sodium citrate- 1.47g/.5L; pH 6.0) for unmasking free floating 40 um sections (boiling on hot plate for 5 min). After unmasking the whole of the sections turn blue black (DAB) as compared to sections whcih are not boiled. Also, with fluorescent method, there is more non specific binding though there is increase in specific binding also for some antigens.
What acan be the reason for increase in non specific binding by boiling the sections?
Regards
human

Citrate buffer antigen unmasking
« on: October 19, 2003, 03:55:57 AM »

Offline ghostdog

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Citrate buffer antigen unmasking
« Reply #1 on: October 20, 2003, 04:49:02 AM »
Are you letting the slides cool down after the boiling , this is a big cause of non-specific binding of DAB

Offline Ghuman

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Citrate buffer antigen unmasking
« Reply #2 on: October 20, 2003, 04:55:34 AM »
I boil free floating sections and let them cool for 3 min and then add cool citrate buffer to decrease the temperature quickly. what should i do is this cooling down ok .
human

Offline ghostdog

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Citrate buffer antigen unmasking
« Reply #3 on: October 21, 2003, 05:22:53 AM »
yeah it sounds ok, have a look at the topic section on antigen retrieval

Offline cheewee

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try tris/edta ph9.0
« Reply #4 on: November 06, 2003, 11:58:04 PM »
I use tris/EDTA, ph9.0 it's give me a better results, why don try?

try tris/edta ph9.0
« Reply #4 on: November 06, 2003, 11:58:04 PM »