Optimization of polymerase chain reaction for detection of HIV type 2 DNA.

Important note: Information in this article was accurate in 1999. The state of the art may have changed since the publication date.

Optimization of polymerase chain reaction for detection of HIV type 2 DNA.

AIDS Res Hum Retroviruses. 1998 Sep 1;14(13):1151-6. Unique Identifier : AIDSLINE MED/98407604
Walther L; Grankvist O; Mirzai B; da Silva Z; Fredlund H; Biberfeld G; Thorstensson R; Swedish Institute for Infectious Disease Control and Microbiology; and Tumor Biology Center, Karolinska Institute, Stockholm.

Abstract: The aim of this study was to increase the sensitivity of an earlier version of an HIV-2 nested PCR assay based on primers in the gag, pol, LTR, and env regions. The assay was first optimized with regard to concentrations of dNTP, MgCl2, and primers, using a method that allowed optimization of all three parameters in only two test runs. We then designed and optimized new primer sets in the LTR, gag, and gag/pol regions that were based on more isolates than were the former (old) primer sets. Samples from 57 HIV-2 antibody-positive individuals were tested with the four old primer sets as well as with the three new primer sets. Five primer sets from this run (new gag, new gag/pol, old LTR, old env, and new LTR) were then tested with 35 more samples, giving a total number of 92 tested samples from HIV-2-infected individuals. At initial testing of the 92 samples a combination of primer sets from two different regions yielded a sensitivity ranging from 93.5 to 98.9%. After repeated testing the sensitivity ranged from 96.7 to 100% for the different primer combinations. The specificity was 100% for all primer sets except old LTR, which had a specificity of 97%. In conclusion, it is possible to create a more sensitive PCR assay by optimizing the different PCR parameters as well as by including primer sets based on more isolates.

Keywords: JOURNAL ARTICLE DNA Primers/GENETICS DNA, Viral/*ANALYSIS Gene Products, gag Human HIV Infections/VIROLOGY HIV Long Terminal Repeat HIV Seropositivity/*VIROLOGY HIV-2/*GENETICS/*ISOLATION & PURIF Polymerase Chain Reaction/*METHODS Sensitivity and Specificity Support, Non-U.S. Gov't
990228
A9920920

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