CLL1141

Sigma-Aldrich

A549 Cells GFP-EGFR

NACRES:
NA.81
Pricing and availability is not currently available.

Quality Level

biological source

human male lung (Source Disease: Lung carcinoma)

OMIM accession no.

storage temp.

−196°C

Gene Information

human ... EGFR(1956), ERBB1(1956)

General description

Glucocorticoid is necessary for A549 cell induction to respond to OSM (oncostatin M), interleukin (IL-6) and fibroblast conditioned medium. The EGFR gene is mapped to human chromosome 7p11.2 and encodes a 170 kDa receptor protein belonging to the HER family of receptor tyrosine kinases. Egfr is localized to epithelial cell surface.
A549 GFP-EGFR are lung carcinoma, epithelial cells from a human 58 year old male caucasion having a ZFN modification creating an EGFR-GFP transgene expressed from the endogenous EGFR gene locus.

This cell line was derived from ATCC catalog No. CCL-185.

Application

A549 Cells GFP-EGFR is an A549 cell line in which the genomic EGFR gene has been endogenously tagged with a Green Fluorescent Protein gene (GFP) using CompoZr® Zinc Finger Nuclease technology. Integration resulted in endogenous expression of the fusion protein in which GFP is attached to the C-terminus of EGFR. Fluorescence imaging shows characteristic EGFR membrane expression. Upon the addition of EGF, the cell line shows redistribution of EGFR from the cell membrane to endosomes, making it useful for high content screening of compounds. For example, the redistribution can be abolished by a selective inhibitor of EGFR, Tyrphostin AG 1478. This stable cell line was expanded from a single clone. The target′s gene regulation and corresponding protein function are preserved in contrast to cell lines with overexpression via an exogenous promoter.

Features and Benefits

Zinc Finger Nuclease (ZFN)-mediated targeted integration of a fluorescent GFP tag into the last coding exon of the EGFR gene on chromosome 7p11.2 to create a cell line exhibiting stable expression of the transgene tagged with GFP on the C-terminus of the protein.

These A549 cells are adherent with a doubling time of approx. 22 hours.

Quality

Tested for Mycoplasma, sterility, post-freeze viability, short terminal repeat (STR) analysis for cell line identification, cytochrome oxidase I (COI) analysis for cell line species confirmation.

Preparation Note

Media Renewal changes two to three times per week.

Rapidly thaw vial by gentle agitation in 37°C water bath (~2 minutes), keeping vial cap out of the water. Decontaminate with 70% ethanol, add 9 mL culture media and centrifuge 125 x g (5-7 minutes). Resuspend in complete culture media and incubate at 37°C in a 5% CO2 atmosphere.

Subculture Ratio: approx. 1:3-1:6

The base medium for this cell line is RPMI-1640, Cat. No. R0883. To make the complete growth medium, add the following components to the base medium: fetal bovine serum, Cat. No. F2442, to a final concentration (v/v) of 10% and L-glutamine, Cat. No. G7513, at a final concentration of 2 mM.

Cell freezing medium-DMSO 1X, Cat. No. C6164.

Legal Information

CompoZr is a registered trademark of Sigma-Aldrich Co. LLC

WGK Germany

WGK 2

Flash Point(F)

192.2 °F - closed cup

Flash Point(C)

89 °C - closed cup

Certificate of Analysis
Certificate of Origin
Kenji Tanabe
Scientific reports, 6, 25095-25095 (2016-04-28)
Small-molecule compounds are widely used as biological research tools and therapeutic drugs. Therefore, uncovering novel targets of these compounds should provide insights that are valuable in both basic and clinical studies. I developed a method for image-based compound profiling by...
Cláudia M Botelho et al.
Journal of biophotonics, 11(9), e201700323-e201700323 (2018-03-15)
Epidermal growth factor receptor (EGFR) plays a key role in regulating cell survival, proliferation and migration, and its overexpression and activation has been correlated with cancer progression. Cancer therapies targeting EGFR have been applied in the clinic with some success....
Culture of Animal Cells: A Manual of Basic Technique and Specialized Applications (2015)
Effect of epidermal growth factor receptor gene polymorphisms on prognosis in glioma patients.
Li B, et al.
Oncotarget, 7(39), 63054-63054 (2016)
Catarina Oliveira Silva et al.
PloS one, 11(10), e0165419-e0165419 (2016-10-28)
The application of functionalized nanocarriers on photothermal therapy for cancer ablation has wide interest. The success of this application depends on the therapeutic efficiency and biocompatibility of the system, but also on the stability and biorecognition of the conjugated protein....

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