# 403: BSK-MEDIUM


## Main sol. 403

1000	ml	**Solution A**	
200	ml	**Solution B**	
143	ml	**Solution C**	
86	ml	**Solution D**	

1. Combine the warm (37°C) solutions A, B, C, D, and filter-sterilize the warm medium. Solution B can also be added after filtration.
2. Alternatively, buy supplemented or un-supplemented BSK-H medium (Fa. Bio&SELL or Sigma) and supplement the medium according to manufacturer's advice. Sterilize the mixed and warm (37°C) medium by filtration.


## Solution A

100	ml	CMRL 1066 (GIBCO 042-1545, glutamin 10 x)	
5	g	Proteose peptone no. 2 (DIFCO 0121-01-3)	
1	g	Tryptone peptone (DIFCO 0123-01)	
1	g	Proteose yeastolate (TC) (DIFCO 5577-15)	
6	g	HEPES buffer (acid) (SIGMA H3375)	
0.7	g	Sodium citrate	
3	g	Glucose (alpha-, D+)	
0.8	g	Na-pyruvate	
0.4	g	N-Acetylglucosamine	
0.3	g	MgCl2 x 6 H2O	
0.1	g	L-Glutamine	
900	ml	Double distilled water (filter-sterilized)	

1. Stir slowly at 4 - 10°C for 3 hours, adjust pH to 7.6 with 5 M NaOH. Filter sterilize.


## Solution B

14	g	Gelatine	
200	ml	Double distilled water	

1. Dissolve 14.0 g gelatine in 200 ml bidest. water, autoclave 15 minutes at 115°C.


## Solution C

50.05	g	Bovine serum albumin (Sigma No A9647)	
143	ml	Double distilled water	

1. Dissolve 50.05 g bovine serum albumine, fract. V (important: Sigma No A9647) in 143 ml bidest. water, or use BSA-solution Sigma A7409. 


## Solution D

86	ml	Rabbit serum (inactivated, GIBCO 037-06120M)