View source for Labelling Pax6 with Dig-NHS

== Procedure ==
Pax6 AB  is at 5 mg/ml, 1 ml

1)	Dialyze 500 µl into PBS O/N then one change in morning 2 hours.

2)	Spin antibody and check protein concentration by BioRad

STD			OD 595

0

1

3

6

9


1 µl Pax6



3)	Bring antibody to approx. 2.5 mg/ml in PBS.  Final vol:

4)	Check 1M NaHCO3 solution for pH (should be pH 8.3)

5)	Add 1/10 volume of 1M NaHCO3.  


6)	Remove some Pax6 AB as non-labelled control.

7)	Thaw NHS-Dig and spin briefly.  
Add Dig-NHS (10 mg/ml stock in DMSO) at 1:50 dilution to AB  
 (20 µl per 1 ml) and mix rapidly
(Dig is stored in liq. Nitrogen, tower 6 box H)

8)	Incubate 1 hour at RT with occasional mixing

9)	Add 100 mM Glycine pH 7   (30 µl per 500 µl of AB)   incubate atleast 10 min.

10)	Dialyze labeled and unlabeled sample into TBS O/N

11)	Store with 0.1% BSA in 50% glycerol at -20 C.