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Counting and measuring IMPs and pits: why accurate counts are exceedingly rare
1Department of Anatomy and Neurobiology, Colorado State University, Ft. Collins 80523.
Journal of Electron Microscopy Technique
|November 1, 1989
Summary
Accurate membrane analysis requires high complementarity between fracture faces. Simple procedural changes and discarding pre-fractured images improve intramembrane particle (IMP) and pit counts for reliable molecular change identification.
Area of Science:
- Cell Biology
- Biophysics
Background:
- Particle counting and measuring are used to study membrane changes.
- Current methods often compromise membrane face complementarity due to prefractures.
Purpose of the Study:
- To address the premature use of particle counting for membrane molecular change identification.
- To establish criteria for reliable intramembrane particle (IMP) analysis.
Main Methods:
- Implementing simple procedural alterations to improve replica quality.
- Recognizing and discarding images with cryogenic or mechanical prefractures.
- Describing a cleaving method to avoid water vapor contamination.
Main Results:
- Achieved high complementarity between membrane faces by discarding non-complementary images.
- Identified equivalent numbers and near-identical diameters of membrane pits and IMPs in complementary faces.
- Demonstrated that optimized cold fracture and immediate replication minimize contamination, yielding more accurate IMP and pit size and number data.
Conclusions:
- Complementarity of membrane faces is crucial for validating IMP counts.
- Perceived changes in IMP density or size should only be accepted when high complementarity is achieved.
- Reliable IMP analysis is essential for understanding developmental, disease, and pharmacological effects on membranes.