intracellular cytokine detection

Timothy L Wyant twyant at umaryland.edu
Tue Mar 10 19:09:15 EST 1998


On Mon, 9 Mar 1998, frederic clement wrote:

> Dear scientists,
> 
> Can anyone help me with commercial available permeabilisation kits or
> other commercial stuff for detection of intracellular cytokines by
> flow-cytometry ? 
> 
> Thanks a lot.
> 
> PS : Please mail me your advice.
> 
> 
Several companies including Pharmingen (San Deigo) and Coulter sell kits
to permeabilize cells.  However, inour laboratory we have found that the
following procedure works quite well.

1)  Surface stain as normal
2) fix with 4%formaldehyde
3) Wash with PBS
4) add PBS containing 0.1% saponin and stain intracellularly for 30 min.
5) Wash out intracellular stain with PBS-azide containing 0.1% saponin.
6) fix with 1% formaldehyde

This is the basic procedure.  The literature has published several methods
based on this type of procedure

Good luck

Dr. Tim Wyant




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