Tris-Tricine protocols

Volker Nölle a2239221 at SMAIL1.RRZ.UNI-KOELN.DE
Thu Mar 13 05:57:11 EST 1997


We use a modified and easy protocol of Tricine-SDS-gels with good results:

16.5 % gel:

4 ml 49.5 % Acryl/Bis (100:3)
4 ml 3 M Tris 0.3 % SDS pH 8.45
1.6 g Glycerol
2.4 ml MQ


4 % stacking gel:

0.5 ml 49.5 % Acryl/Bis (100:3)
1.55 ml Tris 0.3 % SDS pH 8.45
4.2 ml MQ


Anode buffer:
0.2 M Tris pH 8.9

Kathode buffer:
0.1 M Tris pH 8.25
0.1 M Tricine
0.1 % SDS

Run overnight at 12-16 mA in a gel as big as possible (about 20 cm).


Dye with colloidal Coomassie (do not use Destain before !):

0,8 g Coomassie G-250 in 400 ml MQ=20
add 400 ml 1 M H2SO4
stir for 3 h, filtrate (folden filter)
add 88 ml 10 M KOH and 124 ml 100 % TCA

dye for some hours or overnight at room temperature
then 1 or 2 hours in water
sensitivity: 0,2 =B5g protein

good luck!


Volker Noelle
Institute of Biochemistry
University of Cologne
Germany

email: a2239221 at smail.rrz.uni-koeln.de




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