Journal
CYTOMETRY PART A
Volume 71A, Issue 9, Pages 693-699Publisher
WILEY-LISS
DOI: 10.1002/cyto.a.20429
Keywords
calcium pump; flow cytometry; erythrocyte; Fluo4
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Funding
- NHLBI NIH HHS [U54 HL070583] Funding Source: Medline
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Differences among red blood cells in the activity of the plasma membrane Ca2+-ATPase (PMCA) can impact cell signaling and survival. However, no method has been reported that measures this activity directly in individual cells. We have designed a novel assay for PMCA activity that uses the fluorescent Ca2+-reporter Fluo4 and flow cytometric analysis. The method recognizes the extrusion of Ca from the cell after a short Ca loading pulse, which avoids the problem of ATP depletion and ascertains activity at V,na, capacity. Our assay is responsive to known PMCA inhibitors, and while not intended for quantitative kinetic analysis of Ca2+ pumping, it can be used to determine qualitative differences between red blood cell populations that vary in PMCA activity. Using this assay, we confirmed that a normal red blood cell population shows heterogeneity with respect to the PMCA V-max. We report a novel assay of PMCA activity in red blood cells that can provide qualitative information on PMCA activity in individual cells. (c) 2007 international Society for Analytical Cytology.
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