Species: Mouse

Cell type: Mammary carcinoma

Transgenes: Murine sodium iodide symporter (mNIS) with neomycin resistance (Neo) for selection with G418 and enhanced green fluorescent protein (eGFP) with puromycin resistance (Puro) for selection with puromycin

Media: RPMI, 10% FBS, 1% Pen/Strep, 0.1 mg/mL G418, 2 μg/mL puromycin

Description: 4T1-mNIS-Neo/eGFP-Puro is a polyclonal population of the murine mammary carcinoma cell line 4T1 (ATCC® CRL-2539™) transduced with 1) LV-mNIS-P2A-Neo (LV025) encoding murine sodium iodide symporter (mNIS) cDNA under the spleen focus-forming virus (SFFV) promoter linked to the neomycin resistance gene via P2A sequence, and 2) LV-eGFP-PGK-Puro (LV031) encoding enhanced green fluorescent protein (eGFP) cDNA under the spleen focus-forming virus (SFFV) promoter and the puromycin resistance gene under control of the PGK promoter.

The lentiviral vectors used are self-inactivating (SIN) vectors in which the viral enhancer and promoter has been deleted. Transcription inactivation of the LTR in the SIN provirus increases biosafety by preventing mobilization by replication competent viruses and enables regulated expression of the genes from the internal promoters without cis-acting effects of the LTR (Miyoshi et al., J Virol. 1998).

Cell Line Authentication: The parental 4T1 cell line was authenticated and certified free of interspecies cross-contamination by short tandem repeat (STR) profiling with 27 STR loci.

Recommended uses:

In vitro: This is a high mNIS/eGFP expressing cell line suitable for use as a positive control cell line in iodine uptake and fluorescence assays to verify NIS or GFP expression respectively in your lentiviral transduced cells.

In vivo: 4T1 cells form tumors post implantation into immunocompromised or syngenic Balb/c mice. The in vivo growth of these tumors can be monitored using noninvasive, high-resolution 3D PET/SPECT imaging or optical imaging for GFP fluorescence.

Note: In-life imaging for GFP fluorescence is not recommended due to high background autofluorescence. Tissues may be harvested post mortem for analysis by conventional microscopy.

Publications that used associated reagents: 

LV-eGFP-PGK-Puro (LV031): Shen et al. Immunovirotherapy with vesicular stomatitis virus and PD-L1 blockade enhances therapeutic outcome in murine acute myeloid leukemia. Blood. 2016. March 17: 127(11): 1449-58. 

Morphology: Low- and high-density cell morphology (200x)

 

Transgene Validation:

NIS function: NIS mediated I-125 uptake assay in the presence or absence of KClO4, a competitive inhibitor of iodine uptake.

 

Flow Cytometry for eGFP: 4T1-mNIS-Neo/eGFP-Puro (green) or control (4T1-mNIS-Puro; grey) cells were fixed with paraformaldehyde and analyzed by flow cytometry (20,000 events).