View source for Purifying IgG1 from Hybridoma Supernatant

== Materials ==
* Supernatant (100-200 ml)
* Peristaltic pump
* 1 ml HiTrap ProteinG column (binding capacity = 25 ml)
* PBS  (100 ml)
* 10x PBS
* .1 M glycine pH 2.7 (25 ml)
* 1 M Tris pH 9.0  (10 ml)
* 10 Eppendorf tubes
* 20% ETOH/H2O  (25 ml)

== Procedure ==
# Add 0.1 volume 10X PBS to supernatant and filter througn 0.22 µm filter to prevent clogging.  Keep until ready to run on column
# Pump atleast 5 ml of ddH2O through column at 1 ml per minute
# Pump 10 mls PBS through column
# Pass supernatant over the column twice.  (Save supernatant)
# While waiting, prepare 10 labeled tubes for collecting fraction, and put 100 µl of 1M Tris pH 9.0 inside
# Wash column with 15 ml PBS, then stop pump
# Collect 1 ml fractions into the preprepared tubes by eluting in 0.1 M glycine, pH 2.7.  Collect 10 fractions, and mix immediately with Tris to bring pH to 7.0
# Wash column with 10 ml PBS
# Wash column into 10 ml 20% ETOH and store at 4 C.
# Quantitate protein with BioRAD reagent and dialyze into PBS.