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Submembrane Events In Triggerable Cells Studied by Total Internal Reflection Fluorescence Microscopy.

Published online by Cambridge University Press:  02 July 2020

D. Axelrod ,
L.M. Johns ,
E.S. Levitan ,
G.M. Omann  and
R.W. Holz
D. Axelrod
Affiliation:
Dept. of Physics & Biophysics Research Div., Univ. of Michigan, Ann Arbor, MI48109
L.M. Johns
Affiliation:
Dept. of Pharmacology, Univ. of Michigan, Ann Arbor, MI48109
E.S. Levitan
Affiliation:
Dept .of Pharmacology, Univ. of Pittsburgh, Pittsburgh, PA15261
G.M. Omann
Affiliation:
Dept. of Biological Chemistry & Dept. of Surgery, Univ. of Michigan, Ann Arbor, MI48109
R.W. Holz
Affiliation:
Dept. of Pharmacology, Univ. of Michigan, Ann Arbor, MI48109
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Extract

We have studied the dynamics of certain key processes near the plasma membrane inside two types of chemically-triggerable living cells using total internal reflection fluorescence microscopy (TIRFM). In TIRFM, a laser beam is incident upon the cell/glass-substrate interface from the glass side at an angle greater than the critical angle for total internal reflection. This creates an exponentially decaying evanescent field in the cell medium (with a characteristic depth of > 100 nm) capable of exciting fluorescence selectively from the membrane-proximal regions at cell/substrate contacts. Various ways of setting up the optics for such a system are discussed, involving the use of either prisms or very high aperture objectives.

In one application of TIRFM, the motion of adrenalin-containing secretory granules in the immediate submembrane region of chromaffin cells is examined before and after chemical stimulation that causes the granules to release their contents to the cell exterior.

Type
Novel Approaches to Microscopy Of Living Cells
Information
Microscopy and Microanalysis , Volume 5 , Issue S2: Proceedings: Microscopy & Microanalysis '99, Microscopy Society of America 57th Annual Meeting, Microbeam Analysis Society 33rd Annual Meeting, Portland, Oregon August 1-5, 1999 , August 1999 , pp. 1052 - 1053
Copyright
Copyright © Microscopy Society of America

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