Quantitative stability of DNA after extended storage of clinical specimens as determined by real-time PCR

Abstract

Viral DNA stored for extended periods can be amplified by PCR. However, it is unknown whether stored specimens give accurate quantitative results by newer real-time PCR techniques. We therefore compared herpes simplex virus DNA levels in specimens before and after 16 months of storage. The levels of viral DNA remained stable whether the DNA was stored as purified DNA or unextracted DNA in a whole specimen.

FIG. 1.

HSV DNA remains quantitatively stable for real-time PCR when it is stored for 16 months as purified DNA at 4°C. Dacron swab specimens were obtained in April 2000, and HSV DNA was extracted and quantitated by real-time PCR. The purified DNA was then stored for 16 months at 4°C and reanalyzed in August 2001 by a real-time PCR assay identical to the one used in April 2000.

FIG. 2.

HSV DNA remains quantitatively stable for real-time PCR when stored as whole specimens with unextracted DNA for 16 months at −20°C. Dacron swab specimens were obtained in April 2000 and placed in PCR buffer. DNA was extracted from 200 μl of the buffer containing the swab specimen, and the levels of HSV DNA were analyzed by real-time PCR. The remainder of the specimen was stored for 16 months at −20°C until August 2001, and then DNA was extracted from 200 μl of the buffer containing the specimen and the levels were analyzed by a real-time PCR assay identical to the one used in April 2000.

FIG. 3.

The stability of HSV DNA is equivalent for real-time PCR when it is stored for 16 months as either purified DNA at 4°C or specimens with unextracted DNA at −20°C. Specimens were obtained in April 2000 as described in the text. The DNA was extracted and stored in purified form at 4°C, or specimens with unextracted DNA were stored for 16 months at −20°C. In August 2001 DNA was extracted from the frozen specimens and the levels of HSV DNA were compared to those from stored purified DNA by real-time PCR.