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Isolation of epithelial cells from toad bladder.

H J Rodriguez, D W Scholer, M L Purkerson

The American Journal of Physiology

|February 1, 1980

Summary

A new method isolates toad bladder epithelial cells using enzymes and shearing, yielding intact cells with higher metabolic activity than scraped cells. This technique preserves cellular integrity for research.

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Area of Science:

Cell Biology
Physiology

Background:

Toad bladder epithelium is a model for studying ion and water transport.
Previous methods for cell isolation involved mechanical scraping, causing damage.

Purpose of the Study:

To develop a gentler method for isolating toad bladder epithelial cells.
To characterize the isolated cells and compare their function to traditional preparations.

Main Methods:

Isolated toad bladder epithelial cells using ethylenediaminetetraacetic acid, collagenase, DNAse, and shearing forces.
Avoided scraping the mucosal surface to minimize mechanical damage.
Analyzed cell integrity, cell type distribution, nucleic acid and protein content, oxygen consumption, amino acid incorporation, and cyclic nucleotide production.

Main Results:

The new method yielded intact, isolated epithelial cells with minimal clumping.
All three major epithelial cell types were present in the isolated preparation.
Isolated cells exhibited higher rates of oxygen consumption and amino acid incorporation compared to scraped epithelial sheets.
Cyclic nucleotide production in response to vasopressin and carbachol was comparable between isolated cells and scraped sheets.

Conclusions:

This enzymatic and mechanical isolation technique provides intact toad bladder epithelial cells with preserved metabolic functions.
The method offers an improvement over traditional scraping techniques, reducing cellular damage.
The isolated cells are suitable for studying cellular processes and responses to stimuli.

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