Hello! I want to know if someone can help me with this.
We have a sample that we ran in a qPCR using Sybrgreen PCR technology. The result was negative. We have positive and negative controls and they worked fine, both in the amplification curve and in the melting curve.
However, based on the story of the patient, we conducted a nested conventional PCR. In this case the result was positive, which corresponds with the clinical evolution of the patient.
We have repetead 3 times the qPCR and always is negative.
I only know that this patient was receiving antimicrobials, so I wanted to know if a possilibity can be that the antimicrobials are inhibiting the real time PCR but not the nested PCR? Can that be possible??
I know that another option is that the primers are failing to detect the DNA fragment (primers sequences are different from the real time PCR and the nested PCR). However these primers are actually for a conserved region, and have been tested in other parts of the world, so I am actually not sure what can be happening.
Any input or idea will be greatly appreciated.