View source for Pierce BCA Protein Concentration

==Summary==
We use this method measure the amount of protein we have in our unknown samples

===Method===
# *Place samples in order on ice*
# Use the sonicator to break up all the stuff in the cell, free the proteins
# Turn on the machine 
# Clean tip of probe with milli Q water (DI works too), wipe dry
# Set amplitude at 35% intensity
# Put on ear muffs. Place tube in probe, hit red button for 10 seconds (hold the tube in place)
# Place tube on ice
# Wipe probe. Use water to clean between treatments/ controls. Wipe dry
# Clean probe and turn off

===Method for protein assay===
# Use pierce 660 nm protein assay reagent (brown bottle on bench) and BSA pre-diluted standards (known concentrations. Located in the  -20 fridge door, in a box)  
# Use the lysis buffer from the freezer (same as cell culture) as a blank and obtain a 96 well plate (flat bottom)  
# **do duplicated of each solution, loading each solution twice** 
# Load 10 ul of each solution/ sample 
# Start with the blank on the very left side (A1 and B1) ensure there are no bubbles formed 
# Follow with BSA using the lowest concentration first (A2 and B2)  
# Continue with each concentration until done (A3 and B3)... 
# Add proteins the same pattern  
# Place 150 ml of the reagent into an empty box or box lid (make sure its clean) 
# Using a multichannel pipette, quickly add the reagent to each well. The reaction will start right away. Flush each well 10 times to mix 
# Give the reaction about 2 minutes before taking it to the spectrophotometer 
# At the spectrophotometer  
# Place plate in with no cover 
# Open the program 
# Hit file_new 
# Select ‘plate’ 
# enter 660 (wavelength) 
# hit mix and enter the plate 
# Select read normal 
# *Once data pops up record all the numbers in your lab book, and trash plate* 
# Use an excel data sheet which calculates concentrations