Taqman fast virus 1 step qrt pcr kit

Plasma was isolated from undiluted whole blood by Ficoll-based density centrifugation and cryopreserved at -80°C. Plasma viral loads were quantified as previously described [41 ]. Briefly, viral RNA (vRNA) was isolated from plasma samples using the Maxwell Viral Total Nucleic Acid Purification kit (Promega, Madison WI). Next, vRNA was reverse transcribed using the TaqMan Fast Virus 1-Step qRT-PCR kit (Invitrogen) and quantified on a LightCycler 480 instrument (Roche, Indianapolis, IN). Primers and probes are described in [72 (link)].

For animals infected with SIV, plasma viral load was quantified by quantitative reverse transcription-PCR (qPCR) as previously described (90 (link)). In brief, plasma was isolated from whole blood by Ficoll-based density centrifugation. Viral RNA (vRNA) was isolated from cryopreserved plasma samples using the Maxwell Viral Total Nucleic Acid Purification kit (Promega). Then, vRNA was reverse transcribed using the TaqMan Fast Virus 1-Step qRT-PCR kit (Invitrogen) and quantified on a LightCycler 480 instrument (Roche). Primers and probes used for qPCR are described in (91 (link)). qPCR was performed in triplicate for each sample. The limit of detection was established by performing serial dilutions of a known standard.