A-549 VIM RFP
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Cells contain cytomegalovirus (CMV) DNA sequences
Cells contain SV40 sequences
ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.
In lung cancer, Vimentin intermediate filament (IF) proteins have been implicated in many aspects of cancer initiation and progression, including tumorigenesis, epithelial-to-mesenchymal transition (EMT/MET), and the metastatic spread of cancer. Vimentin expression is generally upregulated when the cell is in the mesenchymal relative to the epithelial status. Here, we created a Vim-RFP reporter cell line (CCL-185EMT) using the CRISPR/Cas9 gene editing platform and the parental A549 (CCL-185) non-small cell lung cancer (NSCLC) cell line (a gold standard for studying EMT in cancer metastasis within the lung cancer research community). The created CCL-185EMT cell line harbors a C-terminal red fluorescent protein (RFP) tag on the vimentin gene. This will enable the tracking of the EMT status of cells in vitro by monitoring RFP expression. The integrity of the Vim RFP knock-in has been verified at the genomic, mRNA and protein level for sequence and expression. Functional evaluation of CCL-185EMT shows sensitivity to anti-EMT drugs PP1 and A83-1. This provides the foundation for the high throughput (HTS) identification of new anti-EMT drugs for metastatic NSCLC therapy. The A549 Vim RFP reporter cell line provides a visualizable, convenient and sensitive platform for research on the mechanisms of metastasis in vitro and the development of new antitumor drugs for metastatic NSCLC.
For A-549 VIM RFP (ATCC® CCL-185EMT™) cells culture medium should be:
F-12K Medium (Kaighn's Modification of Ham's F-12 Medium) (ATCC® 30-2004™) supplemented with 10% Fetal Bovine Serum (FBS) (ATCC® 30-2020™) and 10 µg/ml blasticidin (Gibco catalog # A11139-03).
To ensure the highest level of viability, thaw the vial and initiate the culture as soon as possible upon receipt. If upon arrival, continued storage of the frozen culture is necessary, it should be stored in liquid nitrogen vapor phase and not at -70°C. Storage at -70°C will result in loss of viability.
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Thiery JP, Sleeman JP. Complex networks orchestrate epithelial-mesenchymal transitions. Nat Rev Mol Cell Biol 7(2): 131-142, 2006. PubMed: 16493418
Richardson F, et al. The evaluation of E-Cadherin and vimentin as biomarkers of clinical outcomes among patients with non-small cell lung cancer treated with erlotinib as second- or third-line therapy. Anticancer Res 32(2): 537-552, 2012. PubMed: 22287743
Gilles C, et al. Vimentin contributes to human mammary epithelial cell migration. J Cell Sci 112 (Pt 24): 4615-4625, 1999. PubMed: 10574710