saponin
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From: | Gayle Callis <uvsgc@msu.oscs.montana.edu> (by way of histonet) |
To: | histonet <histonet@magicnet.net> |
Reply-To: | |
Content-Type: | text/plain; charset="us-ascii" |
Saponin must be used in all buffers/diluents from blocking to substrate,
HRP or AP, then you need to CLOSE cell membrans by using a buffer rinse
WITHOUT the saponin BEFORE chromagen step and THEN develop the color.
For Cytokine staining on frozens, 0.1% saponin is recommended.
Found this to be fairly harsh on frozen sections,
required very brief and gentle rinse, with a pipette.
Tween 20, we use (to get rid of background) anywhere from 0.05%
to 0.2%, some may use higher concentrations. Find 0.2% works quite well.
We haven't used Triton X100, but saw a protocol (routine IHC staining)
using Sigma's 10% Triton X100 diluted 5 ml/1000 ml buffer, fairly dilute.
Jules Elias book, Immunopathology, A practical approach to diagnosis
has an excellent discussion on detergents.
Hope this helps.
Gayle Callis
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