Minute™ Adipose Tissue Fractionation Kit

In average, total cellular proteins account for less than 2% of adipose tissues. Fractionation of adipose tissue is technically very challenging due to its high lipid and low protein contents. Adipose tissue, especially white adipose tissue (WAT), has been recognized as an important endocrine and inflammation organ in addition to its energy storage function. Fractionation and analysis of proteins from adipose tissues are critical for understanding many physiological/pathological conditions. The water-oil emulsion present in biological sample is notoriously difficult to separate.
Invent Biotechnologies has developed a novel technology to deal with this issue. A porous filter with unique surface property and pre-defined pore size coupled with a specially formulated detergent-free fractionation buffer is the key to fractionate adipose tissue into two fractions: water-soluble protein fraction containing mainly cytosolic proteins and water insoluble fraction containing mainly plasma membrane, and organelles such as mitochondria. The buffers used in this kit are free of primary amine, detergent and reducing agents.

Application
Isolated proteins are compatible with all downstream applications including TMT labeling, enzyme digestion, MS analysis and other applications.

SDS-PAGE profiles of fractionated adipose tissues

A. SDS-PAGE (10%) Profiles of Fractionated Adipose Tissues.
Lane 1, water soluble fraction of porcine WAT; Lane 2, water-insoluble fraction of porcine WAT; Lane 3, water soluble fraction of rat WAT; Lane 4, water-insoluble fraction of rat WAT.
B. Western Blottings of Fractionated Proteins from Rat WAT. Lane 3, water soluble fraction; Lane 4, water in-soluble 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).

Kit components:                                 

• Buffer A: 15 ml
• Buffer B: 15 ml
• 1.5 ml microfuge tube: 20
• Pestles for 1.5 ml tube: 2
• Filter Cartridge with Collection Tubes: 20
• Protein Extraction Powder: 2 g

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