MMP 2/9 zymographic analysis in breast cancer and surrounding
tissues.
The cytosols fromtumorand its corresponding surrounding tissue were
assayed for MMP activity. One percent type B-gelatin solution prepared
in water, 8 % resolving gel is prepared containing 1% B-gelatin
solution. Samples were prepared in MMP dye and loaded in gel to run at
110V for 60 minute. After completion of electrophoresis the gel
wasactivated by washing buffer (as mentioned in the referenced protocol)
and then incubated in zymographic gel development buffer pH 7.4 at 37ÂșC
for 42 hours. Finally the gel was stained in amide black or Coommasie
blue solution for 1 hour at room temperature in a shaker. The gels were
then destained with solution I for 15 to 30 minutes, followed by
destaining solution II for 3 to 5 hours until clear MMP bands appeared.
After washing the destaining solution the gel was incubated in a gel
preservative solution for 15 minutes and finally scanned with a digital
scanner (21)