Transfection with beta-Gal-Vector

Frank Gaunitz cbkga01 at
Tue Nov 16 10:20:45 EST 1993

 am doing transfection experiments in primary cultures from rat liver 
with vectors that carry a lacZ reporter gene. 
Expression levels are determined by a methyl-umbelliferryl assay.
ONPG is not sensitive enough.
Galacto-light doesn't work, because, our extracts kill the enhancer
used in this assay.
My problem now is, that I have very high backround levels of lysosomal
galactosidase activity.
Different pH in the detection reaction does not lead to a proper 
discrimination between lysosomal and transfected (E. coli) activity
of the enzyme.
Does any one have a good idea, how to kill ? the lysosomal activity
or how to enhance E. coli beta-Gal activity.

Frank Gaunitz
Physiologisch-chemisches Institut
Universitaet Tuebingen
Hoppe-Seyler-str. 4
72076 Tuebingen
+49 07071 293356

e-mail: 	cbkga01 at

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