Quantitative PCR analysis of viral nucleic acid is now used by diagnostic laboratories worldwide and is particularly useful for monitoring viral loads in patients to assess the effect of anti-viral therapy and potentially to detect drug-resistant viral strains. In clinical virology, qPCR is commonly used for the detection and quantification of blood-borne viruses, including hepatitis B and C and human immunodeficiency viruses. However, it is also increasingly used to monitor viral pathogens of transplant patients, including Epstein-Barr virus, cytomegalovirus and BK virus. In recent years, our laboratory has used quantitative real-time PCR to investigate BK viral loads in a transplant patient population (see Protocol 14.3). BKV can cause several clinical manifestations in immunocompromised patients including hemorrhagic cystitis and allograft nephropathy. Since adopting qPCR, several limitations of this technology have been identified, including the impact of PCR inhibitors, poor nucleic acid extraction and the effect of sequence variation in PCR primer target sites. Each one of these will impact upon the accuracy of the qPCR results.
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