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Journal of Clinical Microbiology, April 1998, p. 1070-1073, Vol. 36, No. 4
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Optimization of Specimen-Handling Procedures for Accurate Quantitation of Levels of Human Immunodeficiency Virus RNA in Plasma by Reverse Transcriptase PCR

Ruth E. Dickover,1 Steven A. Herman,2 Khaliq Saddiq,1 Deborah Wafer,1 Maryanne Dillon,1 and Yvonne J. Bryson1,*

Department of Pediatrics, UCLA School of Medicine, Los Angeles, California,1 and Roche Molecular Systems, Somerville, New Jersey2

Received 19 June 1997/Returned for modification 27 August 1997/Accepted 20 January 1998

Human immunodeficiency virus type 1 (HIV-1) RNA levels in plasma are currently widely used clinically for prognostication and in monitoring antiretroviral therapy. Accurate and reproducible results are critical for patient management. To determine the effects of specimen collection and handling procedures on quantitative measurement of HIV-1 RNA, we compared anticoagulants and sample processing times. Whole blood was collected from 20 HIV-1-infected patients in EDTA, acid citrate dextrose (ACD), and heparin tubes, aliquoted, and stored at room temperature. Plasma was separated from whole-blood aliquots prepared at <= 1, 3, 6, 24, and 48 h postcollection and then stored at -70°C until use. HIV-1 RNA levels were determined by the AMPLICOR HIV-1 MONITOR assay. Heparinized plasma samples, which were pretreated with heparinase prior to analysis, had the lowest baseline HIV-1 RNA levels. In the first 6 h, HIV-1 RNA levels decreased by 10, 20, and 31% in EDTA, ACD, and heparin tubes, respectively. From 6 to 48 h postcollection, HIV-1 RNA levels decreased in all anticoagulants, albeit at a slower, more consistent rate. Our results indicate that EDTA should be the anticoagulant of choice for plasma HIV-1 RNA measurement by reverse transcriptase PCR, but ACD tubes are acceptable if the plasma is separated within 6 h of blood collection. Caution must be applied in the interpretation of absolute HIV-1 RNA copy number values obtained with suboptimal specimen collection and processing procedures.

* Corresponding author. Mailing address: Department of Pediatrics, UCLA Medical Center, 10833 LeConte Ave., Los Angeles, CA 90095. Phone: (310) 825-9161. Fax: (310) 206-4764. E-mail: YBryson{at}Pediatrics.medsch.UCLA.edu.

Journal of Clinical Microbiology, April 1998, p. 1070-1073, Vol. 36, No. 4
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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Copyright © 1998 by the American Society for Microbiology. All rights reserved.