View source for Preparing Cell Lysates

Materials
RIPA Buffer (for 10mL lysis buffer)
Tris pH7.4 50mM 500uL
Na Deoxycholate 0.25% 250uL
NP-40 1% 1mL
NaCl 150mM 371uL
EDTA 1mM 20uL
NaVO3 1mM 100uL
NaF 1mM 20uL
Protease Inhibitors 1 tab

Basic Protocol
# Stimulate cells if necessary
# Wash cells 2x1mL with ice cold PBS -/- and aspirate
# Add 200uL RIPA buffer and scrape cells
# Pipet into cold eppendorf tubes
# rotate end over end for 30 minutes at 4C to lyse
# Centrifuge 10 min at 13,000 RPM to clarify
# Transfer 150uL of lysate to fresh tube and add 150uL 2XSDS
# Load gel