Sj. Lillard et al., MONITORING EXOCYTOSIS AND RELEASE FROM INDIVIDUAL MAST-CELLS BY CAPILLARY ELECTROPHORESIS WITH LASER-INDUCED NATIVE FLUORESCENCE DETECTION, Analytical chemistry, 68(17), 1996, pp. 2897-2904
The complex temporal evolution of on-column exocytotic release of sero
tonin and proteins from individual rat peritoneal mast cells (RPMCs) w
as monitored by using capillary electrophoresis. Laser-induced native
fluorescence detection with 275-nm excitation was used, and a detectio
n limit of 1.7 amol (S/N = 3; rms) was obtained for serotonin. A physi
ological rurning buffer was used to ensure that the cell remained viab
le throughout. The secretagogue was polymyxin B sulfate (Pmx). Followi
ng the injection of a single mast cell into the capillary, electromigr
ation of Pmx toward and past the cell induced degranulation and releas
e of serotonin. The time course of release was registered in the elect
ropherograms with subsecond resolution. Subsequent introduction of SDS
caused the cell to lyse completely and allowed the residual serotonin
to be quantified. The average amount of serotonin observed per RPMC w
as 1.6 +/- 0.6 fmol; the average percentage of serotonin released was
28 +/- 14%. Events that are consistent with released serotonin from si
ngle submicrometer granules (250 at each) were evident, each of which
contained an average amount of 5.9 +/- 3 amol.