western blot原理，操作 - 图文(6)

50 mM Tris, pH 8.0

2 RIPA buffer (Radio Immuno Precipitation Assay buffer) 150 mM sodium chloride 1.0% NP-40 or Triton X-100 0.5% sodium deoxycholate

0.1% SDS (sodium dodecyl sulphate) 50 mM Tris, pH 8.0

注：The 10% sodium deoxycholate stock solution (5 g into 50 ml) must be protected from light.

3 Tris-HCl buffer 20 mM Tris-HCl, pH 7.5

4 Tris-Triton buffer 10 mM Tris, pH 7.4 100 mM NaCl 1 mM EDTA 1 mM EGTA 1% Triton X-100 10% glycerol 0.1% SDS

0.5% deoxycholate

All four of these buffers will keep at 4oC for several weeks or for up to a year aliquotted and stored at -20oC.

5 TBS 10x (concentrated TBS) 24.23 g Trizma HCl 80.06 g NaCl

Mix in 800 ml ultra pure water. pH to 7.6 with pure HCl. Top up to 1 L. TBST

For 1 L: 100 ml of TBS 10x + 900 ml ultra pure water + 1ml Tween20

6 PBS: 1.16 g Na2HPO4, 0.1 g KCl, 0.1 g K3PO4, 4 g NaCl (500 ml distilled water) pH 7.4

附录2 SDS-PAGE胶的配制

一 30% (W/V) Acrylamide 的配制

1称量Acrylamide290g，Bis10 g下列试剂，置于1 L烧杯中 2向烧杯中加入约600 ml的去离子水，充分搅拌溶解。

3. 加去离子水将溶液定容至1 L，用0.45