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Related Experiment Videos

A simple isolation method for basal-lateral plasma membranes from rat kidney cortex.

V Scalera, Y K Huang, B Hildmann

    Membrane Biochemistry
    |January 1, 1981
    PubMed
    Summary

    This study details a method for isolating basal-lateral membranes using Percoll gradients. The technique effectively enriches (Na+--K+)-ATPase activity and is applicable to pig kidneys.

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    Area of Science:

    • Biochemistry
    • Cell Biology
    • Membrane Biology

    Background:

    • Basal-lateral membranes are crucial for kidney function.
    • Efficient isolation of these membranes is essential for studying their specific roles.

    Purpose of the Study:

    • To develop and validate a method for isolating basal-lateral plasma membranes.
    • To characterize the isolated membranes using enzyme activity markers.

    Main Methods:

    • Separation of basal-lateral membranes from heavy microsomal fractions using a self-orienting Percoll gradient.
    • Centrifugation at 48,000g for 0.5 hours.
    • Assay of (Na+--K+)-ATPase and adenylate cyclase activities.

    Main Results:

    • The isolation procedure resulted in a 20-fold enrichment of (Na+--K+)-ATPase activity, a marker for basal-lateral membranes.

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  • Adenylate cyclase activity in the isolated membranes showed a 6-fold stimulation by parathyrin.
  • The yield of basal-lateral plasma membranes was 5-10% of the initial amount.

    • Conclusions:

    • The Percoll gradient method provides an effective means for isolating enriched basal-lateral plasma membranes.
    • The isolated membranes exhibit hormone-responsive adenylate cyclase activity.
    • This method is adaptable for use with pig kidney tissue.