Product Introduction

RIPA Lysis Buffer is a traditional fast lysis solution for cells and tissues. The protein samples obtained by lysing tissues and cells with RIPA lysis solution can be used for conventional PAGE, Western, immunoprecipitation (immunol precipitation, IP), immunoprecipitation (CO-IP) and other experiments. RIPA lysate has many formulas, which are divided into three types: strong, medium and weak according to its lysis effect. This product is a RIPA strong lysate. Its main components include 50 mM Tris-HCl (pH 7.4), 150 mM NaCl, 1 mM EDTA-2Na, 1% Triton X-100, 1% sodium deoxycholate and 0.1% SDS. This product is suitable for animal or plant tissue and cell samples, as well as fungal or bacterial samples.


Storage and transportation

Transport with wet ice; store at 4°C and protect from light. The validity period is 18 months.

Instructions

Bring your own protease inhibitor. RIPA lysate (strong) needs to be added with protease inhibitors before use, G2006, G2007, G2008, etc. are recommended to prevent protein degradation. The RIPA lysate (strong) mentioned in the following usage methods all means that protease inhibitors have been added.

For tissue samples:

1. The tissue pieces are washed with pre-cooled PBS (G4202 recommended) to remove blood stains, cut into small pieces and placed in a homogenizer.

2. Add 10 times the tissue volume RIPA lysis solution (strong) low temperature homogenate (recommend the high-speed tissue grinder KZ-III-F and KZ-III-FP independently developed and produced by Servicebio). Note that the amount of RIPA Lysis Solution (Strong) can be added at a ratio of about 50 mg of tissue to 1 mL of Lysis Solution. If the content of tissue protein is low, the amount of lysate can be reduced to increase the protein concentration in the crude extract solution.

3. Transfer the homogenate to a 1.5 mL centrifuge tube and shake. In the ice bath for 30 minutes, pipette repeatedly to ensure complete lysis of tissue cells;

4. Centrifuge at 12000 g for 5 min, and collect the supernatant, which is the total protein solution.

For adherent cell samples:

1. Wash the cells 2-3 times with PBS, and thoroughly suck up the remaining liquid for the last time.

2. According to the ratio of 250 μL lysate for cells in each well of the 6-well plate, draw RIPA lysate (strong) into the cell culture plate and bottle, and shake the plate and bottle repeatedly to make the lysate fully contact the cells for 3-5 min.

3. Scrape the cells with a cell scraper and collect them in a centrifuge tube.

4. Centrifuge at 12000 g for 5 min, and collect the supernatant, which is the total protein solution.

For suspension cell samples:

1. Collect cells by centrifugation.

2. Mix the cell liquid with RIPA Lysis Buffer (Strong) according to the ratio of 250 μL lysis buffer per well of the 6-well plate, and shake.

3. In an ice bath for 30 minutes, pipette several times every 10 minutes during which time to ensure that the cells are completely lysed.

4. Centrifuge at 12000 g for 5 min, and collect the supernatant, which is the total protein solution.

For bacterial or fungal samples:

1. Take 1 mL of bacterial suspension, centrifuge to remove the supernatant, wash once with PBS to fully remove the liquid. Vortex to disperse the bacteria as much as possible.

2. Add 100-200 μL of RIPA Lysis Buffer (Strong), vortex gently to mix the bacteria and Lysis Buffer thoroughly.

3. In an ice bath for 10 minutes, pipette several times every 10 minutes to ensure that the bacteria are completely lysed.

4. Centrifuge at 12,000 g for 5 min, and collect the supernatant, which is the total protein solution.


Precautions

1. Tissues or cells may appear sticky when lysed. It can be pipetted repeatedly or vortexed until it is liquid. If it has been thicker, you can add an appropriate amount of lysis solution.

2. This reagent does not contain protease inhibitors, you need to prepare your own protease inhibitors and add them before use. It is recommended to use our company's G2006, G2007, G2008 and other related protease inhibitors.

3. Please wear lab coat and disposable gloves during operation.