ArticleTuberculosis (Edinburgh, Scotland)2013
A novel assay of antimycobacterial activity and phagocytosis by human neutrophils.
Article in Tuberculosis (Edinburgh, Scotland), 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
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Who cites it
7 citing papers in PubMed.
- Evaluation of the innate immune response of caprine neutrophils against Mycobacterium avium subspecies paratuberculosis in vitro.Veterinary research · 2023Article
- Nanoscale clustering of mycobacterial ligands and DC-SIGN host receptors are key determinants for pathogen recognition.Science advances · 2023Article
- Biofilm of Klebsiella pneumoniae minimize phagocytosis and cytokine expression by macrophage cell line.AMB Express · 2022Article
- An In Vivo Model of SeparateInternational journal of molecular sciences · 2022Article
- Differential Effect of Viable Versus Necrotic Neutrophils onFrontiers in immunology · 2018Article
- Brief Report: HIV-1 Infection Impairs CD16 and CD35 Mediated Opsonophagocytosis of Mycobacterium tuberculosis by Human Neutrophils.Journal of acquired immune deficiency syndromes (1999) · 2016Article
- Effect of Antiretroviral Therapy on HIV-mediated Impairment of the Neutrophil Antimycobacterial Response.Annals of the American Thoracic Society · 2015Article
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Authors and funding
8 authors.
Funding
Abstract
Despite abundant evidence that neutrophils arrive early at sites of mycobacterial disease and phagocytose organisms, techniques to assay phagocytosis or killing of mycobacteria by these cells are lacking. Existing assays for measuring the antimycobacterial activity of human leukocytes require cell lysis which introduces new bioactive substances and may be incomplete. They are also time-consuming and carry multiple risks of inaccuracy due to serial dilution and organism clumping. Flow cytometric techniques for measuring phagocytosis of mycobacteria by human cells have failed to adequately address the effects of organism clumping, quenching agents and culture conditions on readouts. Here we present a novel in-tube bioluminescence-based assay of antimycobacterial activity by human neutrophils. The assay yields intuitive results, with improving restriction of mycobacterial bioluminescence as the ratio of cells to organisms increases. We show that lysis of human cells is not required to measure luminescence accurately. We also present a phagocytosis assay in which we have minimised the impact of mycobacterial clumping, investigated the effect of various opsonisation techniques and established the correct usage of trypan blue to identify surface-bound organisms without counting dead cells. The same multiplicity of infection and serum conditions are optimal to demonstrate both internalisation and restriction of mycobacterial growth.
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Registered trials
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