Indirect immunofluorescence was performed as previously described [26 (link),90 (link)]. The primary antibodies used were anti-CD82 (11-559-C100, Exbio Praha, AS, Vestec, Czech Republic) and anti-E-cadherin (sc-8426) followed by appropriate Alexa Fluor 488-conjugated secondary antibodies (Jackson ImmunoResearch labs, Cambridgeshire, UK). Cells were counterstained with DRAQ5 (Invitrogen-Thermo Fisher Scientific) for nuclei visualisation. The stained cells were washed with PBS and mounted on glass slides. Images were collected on a Leica TCS-SP scanning confocal microscope with 63× objective lens.
Roupakia E., Chavdoula E., Karpathiou G., Vatsellas G., Chatzopoulos D., Mela A., Gillette J.M., Kriegsmann K., Kriegsmann M., Batistatou A., Goussia A., Marcu K.B., Karteris E., Klinakis A, & Kolettas E. (2021). Canonical NF-κB Promotes Lung Epithelial Cell Tumour Growth by Downregulating the Metastasis Suppressor CD82 and Enhancing Epithelial-to-Mesenchymal Cell Transition. Cancers, 13(17), 4302.
Corresponding Organization : University of Ioannina
Other organizations :
Academy of Athens, Biomedical Research Foundation of the Academy of Athens, Columbia University Irving Medical Center, University of New Mexico, University Hospital Heidelberg, Heidelberg University, Stony Brook University, Brunel University of London
Primary antibodies used (anti-CD82 and anti-E-cadherin)
dependent variables
Visualization of CD82 and E-cadherin proteins in cells
control variables
Cell staining protocol, including fixation, permeabilization, and blocking steps (as previously described in references 26 and 90)
Mounting of stained cells on glass slides
Imaging using a Leica TCS-SP scanning confocal microscope with 63× objective lens
positive controls
Not explicitly mentioned
negative controls
Not explicitly mentioned
Annotations
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