Per citar aquest document: http://ddd.uab.cat/record/142533
Quantitative Assessment of Mycoplasma Hemadsorption Activity by Flow Cytometry
García Morales, Luis (Institut de Biotecnologia i de Biomedicina 'Vicent Villar Palasí')
González González, Luis (Institut de Biotecnologia i de Biomedicina 'Vicent Villar Palasí')
Costa García, Manuela (Universitat Autònoma de Barcelona. Servei de Cultius Cel·lulars, Producció d’Anticossos i Citometria)
Querol Murillo, Enrique (Institut de Biotecnologia i de Biomedicina "Vicent Villar Palasí")
Piñol Ribas, Jaume (Institut de Biotecnologia i de Biomedicina "Vicent Villar Palasí")

Data: 2014
Resum: A number of adherent mycoplasmas have developed highly complex polar structures that are involved in diverse aspects of the biology of these microorganisms and play a key role as virulence factors by promoting adhesion to host cells in the first stages of infection. Attachment activity of mycoplasma cells has been traditionally investigated by determining their hemadsorption ability to red blood cells and it is a distinctive trait widely examined when characterizing the different mycoplasma species. Despite the fact that protocols to qualitatively determine the hemadsorption or hemagglutination of mycoplasmas are straightforward, current methods when investigating hemadsorption at the quantitative level are expensive and poorly reproducible. By using flow cytometry, we have developed a procedure to quantify rapidly and accurately the hemadsorption activity of mycoplasmas in the presence of SYBR Green I, a vital fluorochrome that stains nucleic acids, allowing to resolve erythrocyte and mycoplasma cells by their different size and fluorescence. This method is very reproducible and permits the kinetic analysis of the obtained data and a precise hemadsorption quantification based on standard binding parameters such as the dissociation constant Kd. The procedure we developed could be easily implemented in a standardized assay to test the hemadsorption activity of the growing number of clinical isolates and mutant strains of different mycoplasma species, providing valuable data about the virulence of these microorganisms.
Nota: Número d'acord de subvenció MICINN/BFU2010-22209-C02-01
Drets: Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original. Creative Commons
Llengua: Anglès
Document: article ; recerca ; publishedVersion
Matèria: Mycoplasma ; Cell staining ; Flow cytometry ; Cell binding ; Adhesins ; Cell binding assay ; Propidium iodide staining ; Staining
Publicat a: PLoS one, Vol. 9 Issue 1 (January 2014) , p. e87500, ISSN 1932-6203

DOI: 10.1371/journal.pone.0087500


9 p, 2.4 MB

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