Monitoring exocytosis with laser-induced native fluorescence: capillary electrophoresis and imaging microscopy

Sheri J. Lillard; Edward S. Yeung; Michael A. McCloskey

doi:10.1117/12.273510

7 May 1997 Monitoring exocytosis with laser-induced native fluorescence: capillary electrophoresis and imaging microscopy

Sheri J. Lillard, Edward S. Yeung, Michael A. McCloskey

Author Affiliations +

Proceedings Volume 2980, Advances in Fluorescence Sensing Technology III; (1997) https://doi.org/10.1117/12.273510
Event: BiOS '97, Part of Photonics West, 1997, San Jose, CA, United States

Abstract

The complex temporal evolution of exocytotic release of serotonin and proteins from individual rat peritoneal mast cells was monitored. Laser-induced native fluorescence with 275- and 305-nm excitation was used to detect the Polymyxin-B sulfate (Pmx) stimulated exocytosis in capillary electrophoresis (CE) and imaging microscopy, respectively. Events are observed that are consistent with released serotonin from single granules (250 aL each). With CE, a detection limit of 1.7 amol (S/N equals 3; rms) was obtained for serotonin. Following the injection of a cell into the capillary, electromigration of Pmx toward and past the cell induced degranulation and release of serotonin. The time course of release was registered in the electropherograms with sub-second resolution. The average amount of serotonin observed per cell was 1.6 +/- 0.6 fmol; the average percentage of serotonin released was 28 +/- 14%. In complementary experiments, exocytosis was monitored temporally and spatially using native fluorescence imagery microscopy. Real time chemical images of serotonin and protein released from individual cells are obtained. The images show that the amount of material released and the time delay of the event varied from cell to cell, and that different regions of a cell behave asynchronously in releasing material.

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Sheri J. Lillard, Edward S. Yeung, and Michael A. McCloskey "Monitoring exocytosis with laser-induced native fluorescence: capillary electrophoresis and imaging microscopy", Proc. SPIE 2980, Advances in Fluorescence Sensing Technology III, (7 May 1997); https://doi.org/10.1117/12.273510

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