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G2038-100ML 100ml IP Lysis Buffer for Immunol Precipitation

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G2038-100ML 100ml IP Lysis Buffer for Immunol Precipitation

Application: Lyse cells or tissues under non-denaturing conditions to prepare protein samples.

Transportation and Storage: Wet ice bag transportation;Store at -20ºC, effective for 12 months.

IP lysate is a lysate which is used to lyse cells or tissues to prepare protein samples under non-denaturing conditions. Protein samples obtained by cleavage of tissues or cells with this lysate can be used in PAGE, Western blot, immunoprecipitation,IP), CO-IP, CHIP (Chromatin immunopre-cipitation), ELISA and other experiments.

The main ingredients 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, and it can also be used for fungal or bacterial samples.

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Technical Specification

Usage:

Protease inhibitors are provided. The IP lysate should be added with protease inhibitors before use. G2006, G2007, G2008, etc., are recommended to prevent protein degradation. The IP lysate mentioned in the following usage means that protease inhibitors have been added.

 

For tissue samples:

  1. The tissue blocks were washed with pre-cooled PBS (recommended G4202) to remove blood stains, and then cut into small pieces and placed in the homogenizer.
  2. Add 10x tissue volume of IP lysis buffer and homogenate at low temperature (It is recommended to use high-speed tissue grinding instrument KZ-III-F and KZ-III-FP, which is independently developed and produced by Servicebio). Note: the amount of IP lysate can be added in a ratio of approximately 50mg of tissue to 1 mL of lysate. If the content of tissue protein is low, the dosage of lysate can be reduced to increase the protein concentration in crude extract solution.
  3. Transfer the homogenate to a 1.5mL centrifuge tube and oscillate. Ice bath for 30 min, during which the pipette was repeatedly blown to ensure the complete lysis of tissue cells at every 10min;
  4. For 12000g samples, it should be centrifuged for 5 min and collect the supernatant, which is the total protein solution.

 

For adherent samples:

  1. Wash the cells with PBS for 2-3 times, and blot the residual liquid thoroughly for the last time.
  2. Pour IP lysate into the cell culture plate and flask according to the ratio of 250μL cell lysate per well of 6-well plate, shake the culture plate and flask repeatedly, and make the lysate fully contact with cells for 3-5 min.
  3. Scrape off the cells with a cell spatula and collect into a centrifuge tube.
  4. Centrifuge at 12000 g for 5 min and collect the supernatant, which is the total protein solution.

 

For suspension cell:

  1. Wash the cells with PBS for 2-3 times, and blot the residual liquid thoroughly for the last time.
  2. Pour IP lysate into the cell culture plate and flask according to the ratio of 250μL cell lysate per well of 6-well plate, shake the culture plate and flask repeatedly, and make the lysate fully contact with cells for 3-5 min.
  3. Scrape off the cells with a cell spatula and collect into a centrifuge tube.
  4. Centrifuge at 12000 g for 5 min and collect the supernatant, which is the total protein solution.

 

For bacterial or fungal samples:

  1.  1 mL of bacterial suspension was taken, the supernatant was removed by centrifugation, and the liquid was washed once with PBS to fully remove the liquid.Vortex to disperse the bacteria as much as possible.
  2. Add 100-200 μl IP lysate, swirl gently to mix the bacteria and lysate well.
  3. Ice bath for 10 min, during which the pipette was repeatedly blown several times every 2 min to ensure complete lysis of the bacteria.
  4. 12000g was centrifuged for 5 min and the supernatant was collected, which was the total protein solution.

Notes:

  1. Thickness may appear during tissue or cell lysis. The pipettor can be repeatedly blown or oscillated by the vortex instrument until it is liquid.If it has been thicker, you can add an appropriate amount of cracking liquid.
  2. This reagent does not contain protease inhibitors. Please prepare your own protease inhibitors and add them before use. G2006, G2007, G2008 and other related protease inhibitors of our company are recommended.
  3. The total protein solution obtained from the pyrolysis of this product is compatible with our BCA protein quantitative detection kit (G2026).
  4. Please wear a lab coat and disposable gloves during operation.
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