IUBio

Help wanted for T-cells assay

linda dgk.oncologie at pobox.ruu.nl
Fri Apr 18 15:46:07 EST 1997


 archee at buffnet.net wrote:

>I am a new technician in an immunology lab.  We do T-cell
>proliferations using thymidine.  Could someone please tell me what the
>MTT assay is?  I am curious.  Thanks.

The MTT assay is a colormetric assay which measures (1) mitochondrial activity 
and (2) cell numbers (assuming that mitochondrial activity is approximately 
the same in the different test wells).  It does NOT measure proliferation.

Water-soluble, yellow MTT is converted by enzymes in mitochondria to 
water-insoluble, blue formazan.  This occurs in living cells (active 
mitochondria) but not in dead cells (no active mitochondria).  The formazan 
can be dissolved in isopropynol/HCL, and the optical density measured by 
(multi-well) spectrophotometer.  More blue = more living cells.

Within certain limits, the MTT (and related colorimetric assays) is an 
accurate, fast means of determining cell number.  The incubation time is about 
3 hrs, and the process time varies depending on the exact chemical used (some 
versions have a water-soluble product, so processing time is minimal).  This 
can be used  in place of a thymidine incorporation assay in some cases, but 
bear in mind that you get rid of the radiation only to play with some very 
carcinogenic, toxic chemicals.

See article:

Mosmann, Tim.  Rapid Colorimetric Assay for Cellular Growth and Survival: 
Application to Proliferation and Cytotoxicity Assays.
J. Immunol. Methods    65: 55-63  (1983)

Good luck!

linda





More information about the Immuno mailing list

Send comments to us at biosci-help [At] net.bio.net