Description

On average, total cellular proteins account for less than 2% of adipose tissue. Adipose tissue, especially white adipose tissue, has been recognized as an important endocrine and inflammation organ in addition to its energy storage function. Fractionation and analysis of proteins from adipose tissue are critical for understanding many physiological/pathological conditions. However, fractionation of adipose tissue is challenging due to its high lipid and low protein contents. The water-oil emulsion present in biological samples is notoriously difficult to separate, but we have developed a cutting-edge technology to deal with this issue: a porous filter with unique surface property and predefined pore size coupling that has a specially formulated detergent-free fractionation buffer. This is the key to fractionating adipose tissue into two: water-soluble protein fraction and water insoluble fraction. The buffers used in this kit are free of primary amine, detergent and reducing agents. Isolated proteins are compatible with all downstream applications including TMT labeling, enzyme digestion, MS analysis and other applications.




Image:


A. SDS-PAGE (10%) profiles of fractionated adipose tissues. Lane 1, water-soluble fraction of porcine WAT; Lane 2, water-insoluble fraction porcine WAT; Lane 3, water-soluble fraction of rat WAT; Lane 4, a water-insoluble fraction of rat WAT.


B. Western blottings of fractionated proteins from rat WAT; Lane 3. water-soluble fraction; Lane 4 water-insoluble fraction. Proteins were separated in 10% SDS-PAGE and probed with following cellular protein marker antibodies: Anti-Na/K ATPase alpha1, a plasma membrane marker (Upstate, clone 464.6), anti-lamin B1, a nuclear envelope marker (ab16048, abcam Cambridge, MA), anti-ubiquinol-cytochrome C reductase core protein (abcam, ab 96333) and GAPDH, a cytosolic marker (Sigma). The specific protein bands were visualized by a substrate Opti-4CN (Bio-RAD).

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