Product Introduction

IP lysate is a lysate that lyses cells or tissues to prepare protein samples under non-denaturing conditions. The protein samples obtained by lysing tissues or cells with this lysis solution can be used in PAGE, western blot, immunoprecipitation (Immunol Precipitation, IP), co-immunoprecipitation (co-IP), ChIP (Chromatin Immunopre-cipitation) and ELISA experiments .

The main components of this product are 25 mM Tris-HCl, 150 mM NaCl, 1 mM EDTA, 1% NP-40 and 5% Triton X-100. It can be applied to animal or plant tissue and cell samples, as well as fungal or bacterial samples.

Instructions

Bring your own protease inhibitor. Protease inhibitors should be added to the IP lysate before use, and G2006, G2007, G2008, etc. are recommended to prevent protein degradation. The IP lysis solution 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 tissue volume IP lysis buffer 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 IP lysate used can be added at a ratio of about 50 mg of tissue to 1 mL of lysate. 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 every 10 minutes to ensure that the tissue cells are completely lysed;

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, pipet the IP lysate 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 the IP lysis buffer at 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 IP Lysis Buffer and vortex gently to mix the bacteria and Lysis Buffer thoroughly.

3. In an ice bath for 10 minutes, pipette several times every 2 minutes to ensure complete lysis of the bacteria.

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. The total protein solution obtained by lysis of this product is compatible with our company's BCA protein quantitative detection kit (G2026).

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