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Journal of Clinical Microbiology, May 2003, p. 2062-2067, Vol. 41, No. 5
0095-1137/03/$08.00+0     DOI: 10.1128/JCM.41.5.2062-2067.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Development and Verification of an Automated Sample Processing Protocol for Quantitation of Human Immunodeficiency Virus Type 1 RNA in Plasma

Brenda G. Lee,1 Kristin R. Fiebelkorn,2,{dagger} Angela M. Caliendo,2 and Frederick S. Nolte2*

Clinical Laboratories, Emory University Hospital,1 Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, Georgia2

Received 8 November 2002/ Returned for modification 30 December 2002/ Accepted 7 February 2003

We developed and verified an automated sample processing protocol for use with the AMPLICOR HIV-1 MONITOR test, version 1.5 (Roche Diagnostics, Indianapolis, Ind.). The automated method uses the MagNA Pure LC instrument and total nucleic acid reagents (Roche Applied Science, Indianapolis, Ind.) to extract human immunodeficiency virus type 1 (HIV-1) RNA from plasma specimens. We compared the HIV-1 load results for a dilution series (1 to 5 nominal log10 copies/ml) and 175 clinical specimens processed by the automated method to those for the same samples processed by the manual methods specified by the manufacturer. The sensitivity, dynamic range, and precision of the viral load assay obtained by automated processing of specimens were similar to those obtained by an ultrasensitive manual processing method. The results were highly correlated (R2, 0.95), and were in close agreement, with a mean difference of 0.09 log10 (standard deviation, 0.292). The limits of agreement were ±0.58 log10 for results for samples processed by both the manual and the automated methods. These performance characteristics were achieved with a smaller sample volume (200 versus 500 µl) and without a high-speed centrifugation step and required only 15 min of labor for a batch of 32 samples. In conclusion, the automated sample preparation protocol can replace both the standard and the ultrasensitive manual methods used with the AMPLICOR HIV-1 MONITOR test and can substantially reduce the labor associated with this test.

* Corresponding author. Mailing address: Clinical Laboratories, Emory University Hospital, Room F145, 1364 Clifton Rd. NE, Atlanta, GA 30322. Phone: (404) 712-7297. Fax: (404) 712-4632. E-mail: fnolte{at}emory.edu.

{dagger} Present address: Department of Pathology, University of Texas Health Science Center at San Antonio, San Antonio, Tex.

Journal of Clinical Microbiology, May 2003, p. 2062-2067, Vol. 41, No. 5
0095-1137/03/$08.00+0     DOI: 10.1128/JCM.41.5.2062-2067.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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