Isolation of Cellular Lipid Droplets: Two Purification Techniques Starting from Yeast Cells and Human Placentas

Jaana Mannik; Alex Meyers; Paul Dalhaimer

doi:10.3791/50981

Isolation of Cellular Lipid Droplets: Two Purification Techniques Starting from Yeast Cells and H...

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Bioengineering

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JoVE Journal Bioengineering

Isolation of Cellular Lipid Droplets: Two Purification Techniques Starting from Yeast Cells and Human Placentas

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09:41 min

April 1, 2014

DOI: 10.3791/50981-v

Jaana Mannik*¹, Alex Meyers*², Paul Dalhaimer^1,2

¹Department of Biochemistry and Cellular and Molecular Biology,University of Tennessee, ²Department of Chemical and Biomolecular Engineering,University of Tennessee

Two techniques for isolating cellular lipid droplets from 1) yeast cells and 2) human placentas are presented. The centerpiece of both procedures is density gradient centrifugation, where the resulting floating layer containing the droplets can be readily visualized by eye, extracted, and quantified by Western Blot analysis for purity.

The overall goal of this procedure is to isolate lipid droplets from tissue. This is accomplished by first dissecting and separating placental tissue. Next, the cells of the olein are homogenized.

Then the organelles are separated by several centrifugation steps. Finally, the floating layer containing the lipid droplets is collected and the lipid drops are characterized. Ultimately, fluorescence microscopy and western blood analysis are used to show the purity of the lipid droplet fraction.

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