Aitken S.C.,University Utrecht |
Bronze M.,University of Witwatersrand |
Wallis C.L.,Lancet Laboratories |
Stuyver L.,Janssen Diagnostics BVBA |
And 8 more authors.
Journal of Clinical Microbiology | Year: 2013
In resource-limited settings (RLS), reverse transcriptase (RT) inhibitors form the backbone of first-line treatment regimens. We have developed a simplified HIV-1 drug resistance genotyping assay targeting the region of RT harboring all major RT inhibitor resistance mutation positions, thus providing all relevant susceptibility data for first-line failures, coupled with minimal cost and labor. The assay comprises a one-step RT-PCR amplification reaction, followed by sequencing using one forward and one reverse primer, generating double-stranded coverage of RT amino acids (aa) 41 to 238. The assay was optimized for all major HIV-1 groupMsubtypes in plasma and dried blood spot (DBS) samples using a panel of reference viruses for HIV-1 subtypes A to D, F to H, and circulating recombinant form 01-AE (CRF01-AE) and applied to 212 clinical plasma samples and 25 DBS samples from HIV-1-infected individuals from Africa and Europe. The assay was subsequently transferred to Uganda and applied locally on clinical plasma samples. All major HIV-1 subtypes could be detected with an analytical sensitivity of 5.00E+3 RNA copies/ml for plasma and DBS. Application of the assay on 212 clinical samples from African subjects comprising subtypes A to D, F to H (rare), CRF01-AE, and CRF02-AG at a viral load (VL) range of 6.71E+2 to 1.00E+7 (median, 1.48E+5) RNA copies/ml was 94.8% (n = 201) successful. Application on clinical samples in Uganda demonstrated a comparable success rate. Genotyping of clinical DBS samples, all subtype C with a VL range of 1.02E+3 to 4.49E+5 (median, 1.42E+4) RNA copies/ml, was 84.0% successful. The described assay greatly reduces hands-on time and the costs required for genotyping and is ideal for use in RLS, as demonstrated in a reference laboratory in Uganda and its successful application on DBS samples. Copyright © 2013, American Society for Microbiology. All Rights Reserved.
Vijgen L.,Johnson and Johnson Ltd |
Verbeeck J.,Catholic University of Leuven |
Van Kerckhove B.,Johnson and Johnson Ltd |
Berke J.M.,Johnson and Johnson Ltd |
And 3 more authors.
Methods in Molecular Biology | Year: 2013
A hepatitis C virus (HCV) replicon-based protease phenotyping assay has been developed that allows determining the susceptibility of a patient's HCV protease sequence to HCV protease inhibitors. In brief, HCV protease sequences amplified from clinical samples are cloned in a transient HCV genotype 1b replicon backbone, containing a luciferase reporter gene. These protease chimeric replicons are replication-competent when electroporated into susceptible Huh7-Lunet cells. Replication can be quantified by measuring the enzymatic activity of the luciferase protein. This assay is reproducible and robust, and has a high overall success rate for determining the phenotypic susceptibility of HCV genotype 1a and 1b patient-derived protease domains to HCV protease inhibitors. In addition, the HCV genotype 1b protease shuttle backbone also supports efficient replication of HCV genotype 4 protease sequences. © 2013 Springer Science+Business Media, LLC.
Janssen Diagnostics Bvba | Date: 2012-12-11
The current invention concerns the identification of B-cell epitopes (as linear peptides) from human polyoma virus proteins and their use in an immune diagnostic assay.
Janssen Diagnostics Bvba | Date: 2015-02-09
The instant disclosure describes a novel genotype and phenotype assay to elucidate and/or evaluate new potential HIV integrase inhibitors, but also currently approved and experimental compounds that target protease, reverse transcriptase, and RNaseH. This assay allows studying linked mutations and mutational patterns that occur under HAART and experimental therapies.
Janssen Diagnostics Bvba | Date: 2014-04-17
JC virus (JCV) is a human polyomavirus that infects the majority of people without apparent symptoms in healthy subjects. A neuropathogenic JCV variant is the causative agent of progressive multifocal leucoencephalopathy (PML), a disorder following lytic infection of oligodendrocytes that mainly manifests itself under immunosuppressive conditions. A hallmark for JCV isolated from PML-brain is the presence of rearrangements in the non-coding control region (NCCR) interspersed between the early and late genes on the viral genome. Such rearrangements are believed to originate from the archetype JC virus variant which is shed in urine by healthy subjects and PML patients. Next generation sequencing (pyro-sequencing) has been performed to explore the NCCR variability in urine of healthy subjects in search for JCV quasispecies and rearrangements reminiscent of PML.