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Fluorescence microscopy study of polymorphonuclear leukocyte substrate attached materials.
J W Francis1, A Y Fabi, H R Petty
1Department of Biological Sciences, Wayne State University, Detroit, Michigan 48202.
Cell Motility and the Cytoskeleton
|January 1, 1988
Summary
This study introduces a new method using DiC18Icc fluorescence to visualize polymorphonuclear leukocyte (PMN) substrate-attached materials (SAMs). This technique reveals complex membrane structures and trailing filopodia during cell locomotion.
Area of Science:
- Cell biology
- Immunology
- Biophysics
Background:
- Substrate-attached materials (SAMs) are crucial for cell function but difficult to visualize.
- Polymorphonuclear leukocytes (PMNs) play key roles in inflammation and immunity.
Purpose of the Study:
- To develop a rapid, non-destructive fluorescence microscopy technique for visualizing PMN SAMs.
- To characterize the structure of PMN SAMs and associated filopodia during locomotion.
Main Methods:
- Incorporation of the fluorescent lipid analog DiC18Icc into living PMN membranes.
- Generation of SAMs by applying shear force (PBS rinsing).
- Visualization of labeled SAMs using fluorescence microscopy without fixation.
Main Results:
- DiC18Icc labeling allows rapid, non-destructive visualization of PMN SAMs.
- PMN SAMs exhibit complex patterns: interconnecting filaments, radiating foci, and smooth areas.
- Trailing filopodia up to 100 microns were observed during locomotion, accounting for ~10% of cell membrane.
Conclusions:
- The DiC18Icc technique provides detailed insights into PMN SAM structure and dynamics.
- Observations support models of cell locomotion involving attachment/detachment steps.
- Data constrain models of membrane flow during leukocyte chemotaxis.