Performance characteristics of the COBAS AmpliScreen HIV‐1 test, version 1.5, an assay designed for screening plasma mini‐pools

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Abstract
BACKGROUND: The COBAS AmpliScreen HIV‐1 test, version 1.5 (v1.5) (Roche Molecular Systems), is designed for screening pools composed of samples from 24 individual units of blood or plasma. A specimen‐processing procedure (Multiprep) simultaneously concentrates and extracts HIV‐1, HCV, and HBV particles from plasma and incorporates an HIV‐1 internal control (IC) RNA. Processed samples are amplified by RT‐PCR using HIV‐1‐specific primers and detected by hybridization of the amplified products to HIV‐1‐ and IC‐specific oligonucleotide probes. STUDY DESIGN AND METHODS: Plasma samples containing known quantities of HIV‐1 were used to evaluate analytical sensitivity and precision and to validate a pool testing algorithm. Analytical specificity was evaluated by adding various viruses and bacteria to HIV‐1‐negative plasma. Seroconversion panels were tested to estimate the window‐period reduction achieved by RNA testing. RESULTS: The analytical sensitivity of the test (concentration that yields ≥95% positive results in a set of replicate tests) was 25 copies of HIV‐1 RNA per mL of pooled plasma. Representative strains from all HIV‐1 group M subtypes were reproducibly detected (>95% positive results) at concentrations of 20 to 200 viral particles per mL. The test did not cross‐react with a set of 31 viral and 5 bacterial isolates, and it yielded negative results on a panel of 500 blood samples from HIV‐1‐seronegative donors. Plasma samples containing abnormally high levels of Hb, albumin, triglycerides, or bilirubin did not interfere with the test. HIV‐1 RNA was detected 2 to 14 days before HIV‐1 antibody and 0 to 28 days before p24 antigen. The test specifically detected pools containing a single positive unit with 2400 HIV‐1 RNA copies per mL and correctly identified the positive unit. CONCLUSION: The COBAS AmpliScreen HIV‐1 test, v1.5, has sufficient sensitivity to detect a single infected unit containing 600 copies of HIV‐1 per mL in a pool with 23 uninfected units and should reduce the window period between infection and seroconversion by at least 2 to 14 days.