Detection of GLUT4-exocytosis by flow cytometry

ZANNI RUIZ EMILIA.; PAVAROTTI MARTIN; MAYORGA LUIS S

Congreso; LVII Annual Meeting of the Argentine Society for Biochemistry and Molecular Biology Research (SAIB); 2021

SAIB - SAMIGE

Flow cytometry is a widely used technique both in medical and scientific fields. It enables the observation of different cellular and molecular parameters at high speed, allowing discrimination between different cell populations. Cell’s glucose uptake requires glucose transporter proteins (GLUTs) on the plasma membrane, where GLUT4 is the main transporter in muscle and adipose cells. Insulin and/or muscle contraction triggers transport and fusion of GLUT4-positive vesicles with the plasma membrane, therefore increasing the number of GLUT4 transporters on the cell surface, a mechanism known as "GLUT4" exocytosis. However, beyond the massive uses of cytometry, it has been sub-employed in the study of GLUT4 exocytosis. Accordingly, our goal was to develop a staining protocol to detect the transporter on the surface of a rat myoblast cell line L6 expressing GLUT4 tagged with myc. Our results show an efficient detection of the GLUT4 increment in insulin-stimulated L6 cells. The results were similar to those obtained by immunofluorescence and ELISA, two assays widely used to study GLUT4 exocytosis. We conclude that flow cytometry is a rapid, quantitative, and precise technique to evaluate GLUT4 exocytosis in thousands of cells.