Uni gold recombigen hiv

The extracted DNA was amplified for HPV using a nested PCR approach with GP5+/GP6+( [GP5 + 5¹-TTT GTT ACT GTG GTA GAT ACTAC-3¹], GP6+[5¹- GAAAAATAAACTGTAAATCATATTC-3¹] ) and PGMY 09/11 primer sets (MY/GP(+) [11 (link)] that amplify a conserved 150 bp sequence of the L1 open reading frame. AccuPowerHotStart PCR Premix (Bioneer Corporation, South Korea) was used for the PCR while genotypic identification was achieved by direct sequencing using the Gp6+ oligoprimer. Assays were normalized to a reference gene and a calibrator was included in every run. Thermal cycler (BioRad) and Beckman Coultier CEQ 2000XL were used for PCR and sequencing respectively. Sequence alignments were performed against various standard HPV genotype sequences stored in the GenBank database by on-line BLAST analysis to arrive at specific genotyping. HIV diagnosis was confirmed in all participants using the standard national testing algorithm of rapid immunoassays: Uni-Gold™ Recombigen^® HIV (Trinity Biotech plc, Bray, Ireland) and Determine^® HIV-1/2 (Abbott Japan co Ltd, MinatoKu, Tokyo, Japan). Indeterminate results were subjected to enzyme-linked immunosorbent assay to confirm HIV status.

Trained counselors used finger prick to conduct HIV rapid tests, following South Sudan’s national HIV testing guidelines at the time; parallel tests used Uni-Gold^™ Recombigen^® HIV (Trinity Biotech, Bray Country, Ireland) and Determine^® HIV 1/2 (Inverness Medical Innovations, Petchabun, Japan) tests. Bioline^® HIV 1/2 3.0 (Standard Diagnostics, Inc., Korea) was used as a tiebreaker for discordant results.
Dried blood spot samples drawn by lancet from the finger were collected for HIV rapid test QC, malaria and VL testing in 2010, and syphilis testing in 2012, and storage for future testing.

The outcome variable HIV serostatus at baseline was assessed with two HIV rapid tests [40 ] done in parallel with the FDA-approved Uni-gold™ Recombigen^® HIV (Trinity Biotech plc, Bray, Co. Wicklow, Ireland) and Determine™ HIV-1/2 (Alere Medical Co.Ltd, Matsudo-shi, Chiba, Japan) test. If both of the HIV rapid tests were non-reactive, no further testing was done. If one or both of the HIV rapid tests was reactive, a CD4 cell count was performed and confirmatory test was performed using an FDA-cleared Western blot test. Further details on HIV testing have been described in the HPTN 068 study protocol and the baseline paper [32 ,33 ]. HSV-2 infection testing was performed using the Herpes Simplex Virus Type 2 IgG ELISA assay (Kalon Biological, LTD Guildford, UK), with an index cut-off of 1.5 normalized optical density units. If the HSV-2 test was positive, no further HSV-2 testing was done at the study site at follow-up visits. HSV-2 results were confirmed retrospectively at the HPTN Laboratory Centre.

A gynaecological examination was performed using a swab. Candida colonization was diagnosed by Gram stain; bacterial vaginosis was scored according to Nugent’s criteria. Infection with Trichomonas vaginalis was diagnosed by PCR, using a validated method which was part of the HPV genotyping essay.
A HIV diagnosis was performed using rapid immunoassays: Uni-Gold™ Recombigen® HIV (Trinity Biotech plc, Bray, Ireland) and Determine® HIV-1/2 (Abbott Japan co Ltd, Minato-Ku, Tokyo, Japan). In the case of indeterminate results, an enzyme-linked immunosorbent assay was used to confirm HIV status.

All three components of voluntary counseling and testing (VCT) for HIV (pre-test counseling, HIV testing and post-test counseling) were performed in a single visit, as practiced in national HIV VCT sites in Kenya. Field workers were trained by the Kenya Association of Professional Counselors and certified by NASCOP to deliver full VCT services. The rapid test DetermineHIV-1/2 (Alere GmbH, Cologne, Germany) was used. Testing was performed by finger prick with a sterile lancet to obtain two drops of blood and the result could be read after 15 minutes. Additionally, all participants received another rapid test (BiolineHIV-1/2, Standard Diagnostic Inc., Kyonggi-do, South Korea). Participants with discordant first and second test results underwent a third rapid, tiebreaker test (Uni-Gold Recombigen HIV, Trinity Biotech PLC, Wicklow, Ireland). Dry blood spots were collected for participants who provided consent for testing but declined to know their HIV results and on every 10^th respondent for quality assurance. Dry blood spots were tested at the Medical Research Institute/Center for Disease Control (KEMRI/CDC) in Kisumu, Kenya.

HPV testing was done as described by Depuydt et al. (2006) Micalessi IM et al. (2012) in an accredited laboratory (ISO certification: ISO15189) [29 (link), 30 (link)]. Briefly, HPV DNA was extracted from exfoliated cervical cells using the standard proteinase K-based digestion protocol, following the manufacturer’s instructions. Cells were incubated with proteinase K solution (100 μg/ml) for 3 h at 55 °C. DNA was then further purified by spin column chromatography. HPV types were determined using a series of real-time PCR reactions with specific primers and TaqMan® (Invitrogen, La Jolla, USA) probes for HR HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66 and 68 IARC (2006). Low risk (LR) HPV types 6 and 67 were also detected. HPV DNA was tested according to de Meijers’ et al. (2009) guidelines for HPV DNA test requirements [31 (link)].
The national recommendation was followed to diagnose HIV and a parallel testing algorithm was used for HIV diagnosis using rapid immunoassays: Uni-Gold™ Recombigen® HIV (Trinity Biotech plc, Bray, Ireland) and Determine® HIV-1/2 (Abbott Japan co Ltd, Minato-Ku, Tokyo, Japan). In the case of indeterminate results, an enzyme-linked immunosorbent assay was used to confirm HIV status.