RT-PCR detection limits - (Oct/04/2005 )
Question about the detection limits and sensitivity of RT-PCR.
I often hear of detection levels down to 10-50 copies of amplicon/reaction.
Is this about the lower limits? Can anyone give me an estimate of that in moles/concentration? Probably assuming an amplicon of around 100 bp.
I think you can't estimate that way. Because it is also depend on your PCR efficiency and assay optimization. By the way you can always determine yourself. If you have you pure RNA stock (know amount), you can carry out a 10 fold serial dilution, let say ranging from 10e9 to 10e1 RNA copy. Then add them into your reaction mix and run a real-time RT-PCR. The lowest concentration that can pick up by your assay is your detection limit.
Sometimes this value can be improve by optimization or using bater product.