Non-radioactive Quantitative RT-PCR

[Steve Hopkins]

>I would like to  
>use RT PCR to quantitate mRNA expression.  Most of the techniques I have  
>seen require the use of radioactivity in the amplification reaction.  Are  
>there any techniques available that a reasonably sensitive but don't  
>require the use of radioactivity? I would appreciate any references you  
>might have on this topic.

There are quite a lot, but the method reported by Alard, P 
et al, BioTechniques, v15, 730-737 looks good and we are currently working on 
a variant. Also see Taniguchi et al J. Immunol. Metods, v169, 101-109, though  
I am worried about the competetive PCR method (see posting by T Miller here, 
21 Jan). These methods are microplate based and get away from the gel scanning 
techniques which are quite widely reported. Perkin Elmer (sorry to advertise) 
will sell you a machine that can quantitatePCR cDNA (at a price!).

Steve Hopkins