H100

Cell experiment:

35SO42- efflux is used as a convenient measure of the activity of the Band 3 anion exchanger (AE). Erythrocytes are suspended into MBS containing Na2SO4 rather than NaCl and incubated at 37°C for 30 min. The cell suspension is then centrifuged (3,000 g, 5 min) and the supernatant removed. The above procedure is repeated twice to ensure the intracellular replacement of Cl- with SO42-. To load the erythrocytes with radiolabel, the packed cells are re-suspended to approximately 50% haematocrit in a solution containing 1 part SO42- MBS and 9 parts of a medium containing (in mM) 300 sucrose and 10 MOPS (pH 7.4, 300 ± 5 mOsm) and placed in a microcentrifuge tube. 10 μCi of 35SO42- is then added and the suspension incubated at 37°C for 1 h. At the end of this period, the cells are then washed four times by centrifugation (10,000 g, 10 s) in ice-cold SO42- MBS. All inhibitors (H100) are dissolved in either DMSO or MBS and are added to the cell suspensions prior to the addition of the radioisotope[1].

References:

[1]. Culliford S1, et al. Specificity of classical and putative Cl(-) transport inhibitors on membrane transport pathways in human erythrocytes. Cell Physiol Biochem. 2003;13(4):181-8.