A neutralization assay for HIV-2 based on measurement of provirus integration by duplex real-time PCR.
Specific, effective and rapid neutralization assays are crucial for the development of an HIV vaccine based on the stimulation of neutralizing antibodies and the development of such an assay for the human immunodeficiency virus-2 (HIV-2) is described. Virus neutralization was measured as the reduction of provirus integration using a duplex real-time PCR with high efficiency (99.4%). This PCR uses primers and a probe specific for the proviral LTR. Amplification and quantitative analysis of the cellular GAPDH gene was carried out in parallel to control for toxic or growth-inhibitory components in the sera. The neutralization assay was used to screen sera from 23 HIV-2 infected patients. 21 sera were able to neutralize HIV-2(60415K), 20 sera neutralized HIV-2(7312A) and 7 sera cross-neutralized HIV-1 IIIB. In contrast, when 14 of these sera were tested in parallel with a conventional neutralization assay based on a p27Gag capture ELISA, only one was found to neutralize HIV-2(60415K) and 11 to neutralize HIV-2(7312A) compared with 12 and 13 sera respectively using the PCR-based assay.
Behrendt
R
R
Robert Koch-Institute, Nordufer 20, D-13353 Berlin, Germany.
Fiebig
U
U
Norley
S
S
Gürtler
L
L
Kurth
R
R
Denner
J
J
D000914:Antibodies, Viral / Q000097:blood; D017931:DNA Primers; D004279:DNA, Viral / Q000097:blood / Q000302:isolation & purification; D005260:Female; D015658:HIV Infections / Q000097:blood / Q000276:immunology* / Q000821:virology; D015498:HIV-2 / Q000235:genetics / Q000276:immunology; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000821:virology; D008297:Male; D009500:Neutralization Tests / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D016662:Virus Integration / Q000235:genetics*
D000914:Antibodies, Viral; D017931:DNA Primers; D004279:DNA, Viral