Get 10x Ripa Buffer

How to make a RIPA lysis buffer solution Measure out 3 mL sodium chloride (5 M), 5 mL Tris-HCl (1 M, pH 8.0), 1 mL nonidet P-40, 5 mL sodium deoxycholate (10 %), 1 mL SDS (10%) and add to a 100 mL Duran bottle. Top up the Duran bottle to 100 mL with ddH2O.

RIPA Buffer enables efficient cell lysis and protein solubilization while avoiding protein degradation and interference with the proteins′ immunoreactivity and biological activity. RIPA Buffer also results in low background in immunoprecipitation and molecular pull-down assays.

10X running buffer 29 g Tris base (240 mM) ... Transfer buffer 2.9 g Tris base (12 mM) ... Separating gels X ml Protogel (X = desired % final) ... Stacking gels 1.3 ml Protogel (4 % final) ... Coomassie blue staining solution 0.1% w/v Coomassie blue R250. ... Destaining solution 30% MeOH. ... Longer destaining solution 5% MeOH.

Procedure for Lysis of Tissues Place the fresh tissue into chilled PBS and rinse several times. ... Add RIPA Lysis Buffer to the tissue at 10:1. ... Homogenize for several minutes at high speed until no tissue chunks remain. Incubate on ice for 30 minutes. Centrifuge at ~10000 x g for 10 minutes.

Dilute 10X RIPA Buffer to a 1X solution using ddH2O....Additional notes: For non-adherent cells, add 400 µl of buffer per 107cells once they have been washed in 1X PBS and pelleted. 1X RIPA Buffer can be used for lysis of tissue samples, although a homogenization step is recommended after adding lysis buffer.

1X RIPA Buffer: 20 mM Tris-HCl (pH 7.5) 150 mM NaCl, 1 mM Na2EDTA 1 mM EGTA 1% NP-40 1% sodium deoxycholate 2.5 mM sodium pyrophosphate 1 mM β-glycerophosphate 1 mM Na3VO4 1 µg/ml leupeptin.

Mix 800 μL of water, 100 μL of RIPA Buffer (10X) and 100 μL of SDS Solution in a microtube. - When a different volume is desired, mix water : RIPA Buffer (10X) : SDS Solution = 8:1:1. - For the preparation of RIPA buffer without SDS, mix water : RIPA buffer (10X) = 9:1. - Store 1X RIPA buffer at -20 °C.

1X RIPA lysis buffer consists of 50 mM Tris HCl, 150 mM NaCl, 1.0% (v/v) NP-40, 0.5% (w/v) Sodium Deoxycholate, 1.0 mM EDTA, 0.1% (w/v) SDS and 0.01% (w/v) sodium azide at a pH of 7.4.