14th International AIDS Conference Barcelona, Spain - July 7-12, 2002

Int Conf AIDS 2002 Jul 7-12; 14:(abstract no. B10199)

Cuypers H, Soriano V, Segondy M, Oosterlaken T, Koppelman M, de Mendoza C, Montes B, van de Wiel P
Sanguin Div. CLB, Amsterdam, The Netherlands

BACKGROUND: A HIV-1 viral load assay (NucliSens EasyQ HIV-1) was developed based on NASBA amplification and real time detection with molecular beacon probes. The performance of this assay was evaluated in routine settings at three different sites.

METHODS: HIV-1 RNA is isolated from plasma using the NucliSens Extractor. As an internal control for the entire procedure a fixed amount of calibrator RNA is added to the sample prior to extraction. Subsequently, HIV-1 and calibrator RNA are amplified using the isothermal NASBA reaction and detected by two distinct molecular beacon probes that are added before start of the amplification. One molecular beacon has a loop complementary to a highly conserved region of the wild-type HIV-1 RNA and the other beacon a loop complementary to the calibrator RNA. Quantitation is based on comparison of the fluorescent curves generated by the two beacons that have been linked to different labels.

RESULTS: A linear dose-response was observed in a range of 25-1,000,000 copies/ml plasma with SDs of about 0.10 log at concentrations above 1000 copies/ml and higher SDs at lower concentrations. Quantitations at all sites were very comparable, demonstrating good inter-lab reproducibility. Representatives of group M subtypes A-J and several group O specimens were detected and quantified comparable to other commercially available assays. Parallel testing of clinical samples showed good correlation with the FDA-approved NucliSens HIV-1 QT (R=0.956). Average time to result for amplification and real-time detection was approximately 90 min. for 48 samples including 30 min hands-on.

CONCLUSIONS: This multicenter trial demonstrates a good performance in combination with high throughput of the real-time NucliSens EasyQ HIV-1 assay in routine settings.

020707
B10199

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