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Journal of Clinical Microbiology, October 1998, p. 2964-2969, Vol. 36, No. 10
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Ultrasensitive Reverse Transcription-PCR Assay for Quantitation of Human Immunodeficiency Virus Type 1 RNA in Plasma

Rita Sun,1 Joanne Ku,1 Harsha Jayakar,1 Jo-Chi Kuo,1 Donald Brambilla,2 Steven Herman,1 Maurice Rosenstraus,1,* and Joanne Spadoro1

Roche Molecular Systems, Inc., Branchburg, New Jersey 08876,1 and New England Research Institutes, Watertown, Massachusetts 021722

Received 4 March 1998/Returned for modification 15 May 1998/Accepted 30 June 1998

With the recent introduction of combination therapy, human immunodeficiency virus type 1 (HIV-1) RNA levels in plasma have been dramatically reduced, frequently to below the limit of quantitation (400 copies/ml of plasma) of the AMPLICOR HIV-1 MONITOR Test (Roche Diagnostic Systems). To achieve enhanced sensitivity of the AMPLICOR HIV-1 MONITOR Test, a modified specimen preparation procedure that allows input of RNA from 10-fold more plasma per amplification reaction was developed. This "ultrasensitive" method allows the accurate quantitation of plasma HIV-1 RNA levels as low as 50 copies/ml. A precision study yielded average within-run and between-run coefficients of variation (CV) of 24.8 and 9.6%, respectively. A multicenter reproducibility study demonstrated that the laboratory-to-laboratory reproducibility of this assay is good, with an average CV of 32%. The linear range of this test is between 50 and 50,000 copies/ml of plasma. RNA concentrations measured by the ultrasensitive and standard HIV-1 MONITOR tests exhibited good agreement within the shared linear range of the two methods. The two measurements were within a factor of 2 for 91% of the specimens tested, with the concentration measured by the ultrasensitive method being only slightly lower (median, 22% lower). Preliminary studies suggest that this assay will prove to be useful for predicting the stability of viral suppression in patients whose RNA levels drop below 400 copies/ml in response to highly active antiretroviral therapy.

* Corresponding author. Mailing address: Roche Molecular Systems, 1080 Route 202, Somerville, NJ 08876. Phone: (908) 253-7463. Fax: (908) 253-3318. E-mail: maurice.rosenstraus{at}roche.com.

Journal of Clinical Microbiology, October 1998, p. 2964-2969, Vol. 36, No. 10
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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