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Fig. 5 | BMC Cancer

Fig. 5

From: Cancer-associated mutations reveal a novel role for EpCAM as an inhibitor of cathepsin-L and tumor cell invasion

Fig. 5

Cancer associated EpCAM mutations prevents EpCAM-CTSL localization, interaction and Cathepsin L mediated invasion. a MDCK cells were transduced with WT, M115T, C66Y, and L240A EpCAM and surface expression was analyzed by flow cytometry or confocal microscopy, respectively. b GFP-tagged EpCAM mutants were transfected in 12 well culture plates into HEK-293 T cells. 500uL of Opti-mem media was added to harvest secreted EpCAM in conditioned media after 48 h and assayed by ELISA to measure soluble EpCAM. c After 48 h of culturing, 1-mL of conditioned media was collected from 6-well SKOV3 culture as a source of soluble CTSL. Conditioned media from transformed HEK-293 T cells was used as a source of soluble EpCAM. CTSL activity was determined as discussed in methods. CTSL activity for secreted/cell surface EpCAM was normalized with EpCAM whole cell extracts blot as shown in supplementary Fig. S4B. A5-CM (A549 conditioned media serves as a control). d A549 cells were plated on Matrigel for 6 h. Conditioned media and soluble EpCAM were added, followed by soluble CTSL. After 24 h cells were tabulated to measure invasion

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