Flow cytometric HLA-B27 screening: Cross-reactivity patterns of commercially available anti-HLA-B27 monoclonal antibodies with other HLA-B antigens

Abstract HLA-B27 is a strong diagnostic biomarker as it is found in 90– 95% of ankylosing spondylitis. Routinely, the false-positive results generated by cross-reactivity of HLA-B27 monoclonal antibodies (mAbs) with some other type of HLA-B antigens are problematic. The present study aims at improving the sensitivity and specificity of typing for HLA-B27 by using a more accurate DNA assay. A real time sequence specific primer polymerase chain reaction (PCR-SSP) is performed by using MGB TaqMan oligoprobe and an ABI PRISM™ 7500 sequence detection system. The TaqMan PCR-SSP assay is compared with Immunophenotyping by flow cytometric antigen assay (BD HLA-B27-Kit, clone GS145.2). The results are finally confirmed by sequencing (ABI PRISM ® 3100). As summarised in the table, three methods are identical for clearly positive and negative results. There is 90% concordance for borderline negative results obtained by immunophenotyping with TaqMan PCR-SSC and/or sequencing. However, only 15.4% of borderline positives obtained by immunophenotyping are confirmed as true HLA-B27 positives by sequencing as well as TaqMan PCR-SSC. TaqMan PCR-SSP DNA assay completely correlates with sequencing (gold standard) with 100% PPV and NPV, compared to a PPV of 15% and a NPV of 98% for borderline results by immunophenotyping. Conclusion: when compared with flow cytometric antigen assay, typing HLA-B27 by TaqMan PCR-SSP clearly demonstrates an advantage to better establish the genotype of the patient. Specifically, there is a marked difference for ambitious positive results from immunophenotyping. Moreover, compared to immunophenotyping, there is no need of fresh samples, can test a larger number of samples concomitantly, and reduces technical time as well as cost. N = 52 Immunophenotyping TaqMan PCR-SSP Sequencing Immuno vs Sequencing TaqMan PCR-SSP vs Sequencing Positive 11 11 11 100% 100% Negative 17 17 17 100% 100% Positive (borderline by Immunophenotyping) 13 2 2 15% 100% Negative (borderline ) by Immunophenotyping) 10 9 9 98% 100%

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Flow cytometric analysis on cross-reactivity of human-specific CD monoclonal antibodies with splenocytes of Aotus nancymaae, a non-human primate model for biomedical research

Veterinary Immunology and Immunopathology ◽

10.1016/j.vetimm.2007.06.010 ◽

2007 ◽

Vol 119 (1-2) ◽

pp. 14-20 ◽

Cited By ~ 4

Author(s):

Claudia A. Daubenberger ◽

Rolf Spirig ◽

Manuel E. Patarroyo ◽

Gerd Pluschke

Keyword(s):

Monoclonal Antibodies ◽

Biomedical Research ◽

Flow Cytometric Analysis ◽

Cross Reactivity ◽

Primate Model ◽

Flow Cytometric ◽

Human Primate ◽

Human Specific

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Development of a Colloidal Gold Immunochromatographic Assay for Duck Enteritis Virus Detection Using Monoclonal Antibodies

Pathogens ◽

10.3390/pathogens10030365 ◽

2021 ◽

Vol 10 (3) ◽

pp. 365

Author(s):

Fengli Liu ◽

Yanxin Cao ◽

Maokai Yan ◽

Mengxu Sun ◽

Qingshui Zhang ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Colloidal Gold ◽

Preventive Intervention ◽

Virus Detection ◽

Cross Reactivity ◽

Duck Enteritis Virus ◽

Detection Sensitivity ◽

Immunochromatographic Assay ◽

Efficient Detection ◽

Enteritis Virus

Duck viral enteritis is a highly contagious and fatal disease of commercial waterfowl flocks. The disease occurs sporadically or epizootically in mainland China due to insufficient vaccinations. Early and rapid diagnosis is important for preventive intervention and the control of epizootic events in clinical settings. In this study, we generated two monoclonal antibodies (MAbs) that specifically recognized the duck enteritis virus (DEV) envelope glycoprotein B and tegument protein UL47, respectively. Using these MAbs, a colloidal gold-based immunochromatographic assay (ICA) was developed for the efficient detection of DEV antigens within 15 min. Our results showed that the detection limit of the developed ICA strip was 2.52 × 103 TCID50/mL for the virus infected cell culture suspension with no cross-reactivity with other pathogenic viruses commonly encountered in commercially raised waterfowl. Using samples from experimentally infected ducks, we demonstrated that the ICA detected the virus in cloacal swab samples on day three post-infection, demonstrating an 80% concordance with the PCR. For tissue homogenates from ducks succumbing to infection, the detection sensitivity was 100%. The efficient and specific detection by this ICA test provides a valuable, convenient, easy to use and rapid diagnostic tool for DVE under both laboratory and field conditions.

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Monoclonal antibodies to bovine UDP-galactosyltransferase. Characterization, cross-reactivity, and utilization as structural probes.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(19)57498-1 ◽

1986 ◽

Vol 261 (17) ◽

pp. 7975-7981

Author(s):

J T Ulrich ◽

J R Schenck ◽

H G Rittenhouse ◽

N L Shaper ◽

J H Shaper

Keyword(s):

Monoclonal Antibodies ◽

Cross Reactivity ◽

Structural Probes

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Screening and development of monoclonal antibodies for identification of ferret T follicular helper cells

Scientific Reports ◽

10.1038/s41598-021-81389-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Wenbo Jiang ◽

Julius Wong ◽

Hyon-Xhi Tan ◽

Hannah G. Kelly ◽

Paul G. Whitney ◽

...

Keyword(s):

Animal Model ◽

Monoclonal Antibodies ◽

Lymph Node ◽

Cross Reactivity ◽

Tfh Cells ◽

Follicular Helper ◽

Lymph Node Cells ◽

Human Viruses ◽

Humoral Responses ◽

Murine Monoclonal Antibodies

AbstractThe ferret is a key animal model for investigating the pathogenicity and transmissibility of important human viruses, and for the pre‐clinical assessment of vaccines. However, relatively little is known about the ferret immune system, due in part to a paucity of ferret‐reactive reagents. In particular, T follicular helper (Tfh) cells are critical in the generation of effective humoral responses in humans, mice and other animal models but to date it has not been possible to identify Tfh in ferrets. Here, we describe the screening and development of ferret-reactive BCL6, CXCR5 and PD-1 monoclonal antibodies. We found two commercial anti-BCL6 antibodies (clone K112-91 and clone IG191E/A8) had cross-reactivity with lymph node cells from influenza-infected ferrets. We next developed two murine monoclonal antibodies against ferret CXCR5 (clone feX5-C05) and PD-1 (clone fePD-CL1) using a single B cell PCR-based method. We were able to clearly identify Tfh cells in lymph nodes from influenza infected ferrets using these antibodies. The development of ferret Tfh marker antibodies and the identification of ferret Tfh cells will assist the evaluation of vaccine-induced Tfh responses in the ferret model and the design of novel vaccines against the infection of influenza and other viruses, including SARS-CoV2.

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