Boster Immunoleader

other

Mitochondria Extraction Reagent

AR0156

1 kit

55 USD

AR0156

Mitochondria Extraction Reagent

1 kit

55 USD

Western Blotting Related Reagent

At 4˚C for frequent use, at -20 ºC for long time use.

The Mitochondria Isolation Kit was used to isolate the intact and purified mitochondria from animal cells and tissues. It was used to prepare the mitochondria protein of animal tissues and cultured cells.The kit's procedure is based on Boster’s unique technology that combines high yield and purity with integrity of the isolated mitochondria within 60 min. After lysing with the mitochondria lysate solution (kit component), the obtained mitochondria can be used for SDS-PAGE, Western Blotting, Dimensional electrophoresis and Co-Immunoprecipitation analysis.

Ⅰ. Mitochondria Isolation 1. Sample Preparation a. Tissues homogenate Wash 100-200 mg of fresh tissues twice with 2-4 ml of PBS or physiological saline. Carefully remove and discard the PBS wash, and blot up with filter paper. Cut tissues into small pieces and put them into a small glad homogenizer, add 1-2.5ml of pre-cooling Mitochondria Isolation Kit Reagent A, vortex at medium speed for 10-30 times for homogenate, or incubate for 8-10 min, or use ultrasound method. b. Cultured cells homogenate Digest cells, wash with PBS buffer and centrifuge at 800g for 5-10 min, then collect cells and calculate. Each isolation need at least 5x107 cells, add 1-2.5 ml of pre-cooling Mitochondria Isolation Kit Reagent A, resuspend cells, vortex at medium speed for 10-30 times for homogenate, or incubate for 8-10 min, or use ultrasound method. 2. Transfer tissues or cultured cells homogenate to the centrifuge tube, centrifuge at 800 g for 5 min at 4˚C. Discard precipitate and transfer supernatant to a new 2 ml tube, Centrifuge supernatant at 800 g for 5 min at 4˚C. 3. Centrifuge supernatant at 12,000 x g for 10 min at 4˚C. 4. Remove the supernatant from the mitochondrial precipitate. If desired save the supernatant (cytosolic fraction) for analysis. Ⅱ. Protein Collection 1. Before use, mix Mitochondria Isolation Reagent B and C at 1:10 (i.e. add 10uL Mitochondria Isolation Reagent C to 1ml pre-cooling Mitochondria Isolation Reagent B) and store on ice for use. 2. Add the above solution to mitochondrial precipitate at 10:1 (i.e. add 100uL the above solution into 10uL mitochondrial precipitate. 3. Vortex stroke for 15-30 min at 4˚C. 4. Centrifuge at 10,000rpm for 15 min at 4˚C. Collect the supernatant, deproteinized extractives remains, then albuminimetry. 5. Sub package and store at -70˚C. Avoid repeat frozen and melt.

Used to isolate the intact and purified mitochondria from animal cells and tissues, and prepare the mitochondria protein of animal tissues and cultured cells.

process about 10g tissue or centrifuge and precipitate about 20ml cells