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Adipose tissue especially white adipose tissue (WAT) has been recognized as an important endocrine and

inflammation organ in addition to its energy storage function. Isolation and analysis of proteins from adipose

tissues are increasingly critical for understanding many physiological/pathological conditions. However

isolation and analysis of WAT and brown adipose tissue (BAT) are technically very challengingdue to their

high lipid and low protein contents. The water-oil emulsion present in biological sample is notoriously

difficult to separate. Our company has developed a novel technology to deal with this issue. A porous filter

with unique surface property and pre-defined pore size and thickness coupling with a specially formulated

detergent-free extraction buffer is employed to rapidly and effective separate water-oil emulsion derived from

adipose tissue homogenate. The extraction buffer has lower freezing pointthan that of oil in adipose tissues

and the aqueous phase can be quickly separated from oil phase by passing the tissue homogenate through

the filter. The total proteins isolated are un-biased representation of cellular proteins in the tissue. The

extracted proteins concentration is very high (2-3 mg/ml) as compared to other methods.

Images of water oil enmulsion of porcine WAT before and after passing through the filter

A.SDS-PAGE (10%) profiles of total protein extracted from different adipose tissues. Lane 1, porcine WAT;

Lane 2, chicken BAT; Lane 3, rat WAT.

B. Western blottings of extracted proteins from rat WAT.Proteins were separated in 8-16% gradient 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).