Why is the IC not detectable although the analytical PCR is negative?
FAQ ID -1506

A failure of the IC amplification in the absence of an analytical PCR signal is indicative for either a PCR inhibition or a significant DNA/RNA loss during the nucleic acid isolation procedure (provided the IC was added to the extraction).

Various reasons could account for a failure of IC amplification.

Inhibition of PCR. PCR inhibitors were not quantitatively removed during DNA/RNA extraction. This may particularly occur with stool or urine as sample materials which contain a large number of partially unknown potential PCR inhibitors. Transfer of such molecules may impair or eliminate the DNA polymerase activity. Alternatively, the incomplete removal of ethanol (as a component of washing buffers of many extraction protocols such as the QIAGEN column-based extraction systems) may have caused PCR inhibition. If QIAGEN extraction systems are used, this can be prevented by performing an additional centrifugation step prior to the final elution step (please refer to the artus® user manuals for further details).

Loss of IC during extraction. The use of a non-recommended extraction kit or non-compliance with the extraction kit protocol may lead to loss of DNA/RNA and, consequently, of the IC – provided the IC was added to the extraction procedure. This risk of DNA/RNA loss can significantly be decreased by adding carrier RNA to the extraction. If carrier RNA is not supplied with the extraction kit, QIAGEN recommends the use of homopolymer poly(A) RNA (Amersham Biosciences, cat. no.: 27-4110-01).

Addition of IC to extraction prior to initial lysis step. It is important that the IC is added only after the sample material has been combined with the lysis buffer of the extraction system. If the IC is added prior to this lysis step it may be digested by DNases/RNases present in the sample material. The lysis buffer contains protein-denaturing agents which inactivate all DNases/RNases promptly.