Purification of caveolae by affinity two-phase partitioning using biotinylated antibodies and NeutrAvidin-dextran

Analytical Biochemistry
Irene Barinaga-Rementeria RamírezB Jergil

Abstract

A new concept for affinity two-phase partitioning was tested. The partitioning was based on the interaction of target membranes with a primary antibody which, in turn, interacted with a biotinylated secondary antibody and NeutrAvidin-dextran in a poly(ethylene glycol)/dextran two-phase system. Caveolae selectively redistributed from the top phase to the NeutrAvidin-dextran-containing bottom phase by employing anti-caveolin as the primary antibody. This immunoaffinity approach was more selective than the established sucrose gradient centrifugation method and resulted in highly purified caveolae from Triton X-100-treated liver and lung plasma membranes. The same approach, employing other selective primary antibodies, should facilitate the purification also of other membrane fractions.

References

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Citations

Jul 27, 2007·Proteomics·Djuro Josic, James G Clifton
Sep 15, 2006·Proteomics·Jens Schindler, Hans Gerd Nothwang
May 2, 2006·Journal of Chromatography. a·Henrik EverbergIrene Barinaga-Rementeria Ramírez
Apr 12, 2015·Biophysical Journal·Meaghan E WardVladimir Ladizhansky

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