Immunological detection of pyrazine-2-carboxylic acid for the detection of pyrazinamide resistance in Mycobacterium tuberculosis
Autor: | Elisa Roncal, Edgar A. Florentini, Patricia Sheen, Roberto Alcántara, Daniela E. Kirwan, Ricardo Antiparra, Emily Toscano, Mirko Zimic, Katherine Vallejos, Robert H. Gilman, Noelia Angulo |
---|---|
Jazyk: | angličtina |
Rok vydání: | 2020 |
Předmět: |
Bacterial Diseases
Immunoconjugates Physiology Metabolite Antitubercular Agents Biochemistry Immunologic Adjuvants chemistry.chemical_compound Medical Conditions Immune Physiology Medicine and Health Sciences Public and Occupational Health Enzyme-Linked Immunoassays Bovine serum albumin Enzyme-linked immunoassays 0303 health sciences Immune System Proteins Multidisciplinary biology Chemical Synthesis Serum Albumin Bovine Vaccination and Immunization 3. Good health Actinobacteria Infectious Diseases Medicine Rabbits Research Article medicine.drug endocrine system Science Immunology Enzyme-Linked Immunosorbent Assay Research and Analysis Methods Antibodies Mycobacterium tuberculosis 03 medical and health sciences Inhibitory Concentration 50 Pyrazinoic acid Drug Resistance Bacterial Toxicity Tests Genetics medicine Animals Tuberculosis Chemical synthesis Antigens Immunoassays IC50 030304 developmental biology Detection limit Chromatography Bacteria 030306 microbiology Organisms Immunologic adjuvants Biology and Life Sciences Proteins Correction Pyrazinamide Tropical Diseases biology.organism_classification chemistry Polyclonal antibodies purl.org/pe-repo/ocde/ford#3.02.07 [https] Mutation Immunologic Techniques biology.protein Preventive Medicine |
Zdroj: | PLoS ONE PLoS ONE, Vol 15, Iss 11, p e0241600 (2020) |
ISSN: | 1932-6203 |
Popis: | Pyrazinamide (PZA) susceptibility testing in Mycobacterium tuberculosis (Mtb) is a current area of development and PZA-resistant strains are increasingly prevalent. Previous studies have demonstrated that the detection of pyrazinoic acid (POA), the metabolite produced by the deamidation of PZA, is a good predictor for PZA resistance since a resistant strain would not convert PZA into POA at a critical required rate, whereas a susceptible strain will do, expelling POA to the extracellular environment at a certain rate, and allowing for quantification of this accumulated analyte. In order to quantify POA, an indirect competitive ELISA (icELISA) test using hyperimmune polyclonal rabbit serum against POA was developed: for this purpose, pure POA was first covalently linked to the highly immunogenic Keyhole Limpet Hemocyanine, and inoculated in rabbits. A construct made of bovine serum albumin (BSA) linked to pure POA and fixed at the bottom of wells was used as a competitor against spiked samples and liquid Mtb culture supernatants. When spiked samples (commercial POA alone) were analyzed, the half maximal inhibitory concentration (IC50) was 1.16 mg/mL, the limit of detection 200 μg/mL and the assay was specific (it did not detect PZA, IC50 > 20 mg/mL). However, culture supernatants (7H9-OADC-PANTA medium) disrupted the competition and a proper icELISA curve was not obtainable. We consider that, although we have shown that it is feasible to induce antibodies against POA, matrix effects could damage its analytical usefulness; multiple, upcoming ways to solve this obstacle are suggested. |
Databáze: | OpenAIRE |
Externí odkaz: |