[CMV DNA detection in plasma using real-time PCR based on the SYBR-Green I dye method].

OBJECTIVE

The aim of this study is to assess a real-time PCR technique on the LightCycler 2.0 with SYBR-Green I detection as compared to another real-time PCR method based on detection with FRET (fluorescence resonance energy transfer) probes for the quantification of CMV DNA.

METHODS

The two real-time PCR methods were used to test plasma samples from immunocompromised patients with clinically suspected CMV disease, patients under follow-up without symptoms, and healthy adults. A standard curve for quantitative analysis by the SYBR-Green I method was performed with 10-fold diluted solutions of DNA from the CMV Towne strain (ATCC VR-977) cultured in MRC-5 monolayer. In addition, frozen samples from patients positive for CMV pp65 antigenemia were also analyzed and results compared using the two real time PCR methods.

RESULTS

The real-time PCR technique using SYBR-Green I on the LightCycler 2.0 was a highly specific, fast, simple and reliable test to quantify CMV; moreover, it was cost-effective.

CONCLUSION

Quantification of CMV DNA in plasma using this sensitive, fast, low-cost method was advantageous for the diagnosis and follow up of patients with opportunistic CMV infection, which are increasingly more frequent in our daily hospital clinical practice.