Fig. 3

Validation of qPCR and conventional RT-PCR data by Western blot and zymography. a Western blot analysis: CCL2 protein expression was only detected in TNF-α-treated (100 ng/ml) MDA-MB-435-pFDR1 cells, whereas CLDN1 expression was absent from these cells but was clearly detectable in M13SV1-Cre cells. Increased ICAM1 expression was observed in TNF-α-treated cells, whereas increased MMP9 expression was only found in TNF-α-stimulated M13SV1-Cre cells. Two discrete bands were detected for MMP9 in Western blot analysis; the upper band represents inactive MMP9 and the lower band represents active MMP9. ICAM1 Western blot data were rearranged because ICAM1 samples were originally loaded in a different order. b Zymography: markedly increased MMP9 expression levels and activity were detected in TNF-α-treated M13SV1-Cre cells, whereas even in TNF-α-stimulated MDA-MB-435 cells slightly enhanced MMP9 expression levels could be identified. The data shown are representative of at least three independent experiments