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Giardia/Crypto real time PCR late amplification - (Apr/12/2012 )

I have been running a lot of rtPCR with my bio-rad CFX96. I can not for the life of my figure out why my amplification starts soo late. Im talking about 20-25 cycles. I clean up the DNA with a Norgen kit, then Nanodrop it, get roughly 10-25 ng/nL with 1.65-1.8 260/280 ratios. SsoFast master mix, and primers that we got from other articles. If it is normal to get this late of this amplification, then that would clear it up, if not, can someone please help!


The amplification depends on the amount of template. Is the sample 100% Giardia genomic DNA or you trying to detect the parasite inside other DNA?
It's hard to say without having a positive control with known amount.