Species: Mouse

Cell type: Colorectal Cancer

Transgenes: Near infrared fluorescent protein (iRFP; ex/em = 690/713 nm) with neomycin resistance (Neo) for selection with G418 and murine sodium iodide symporter (mNIS) with puromycin resistance (Puro) for selection with puromycin.

Media: DMEM, 10% FBS, 1% Pen/Strep, 0.4 mg/mL G418, 3 μg/mL puromycin

Description:

CT26.WT-iRFP-Neo/mNIS-Puro is a polyclonal population of the colorectal carcinoma cell line CT26.WT (ATCC® CRL-2638™) transduced with 1) LV-iRFP-P2A-Neo (LV033) encoding the near-infrared fluorescent protein (iRFP) cDNA under the spleen focus-forming virus (SFFV) promoter linked to the neomycin resistance gene (Neo) via a P2A cleavage peptide and 2) LV-mNIS-PGK-Puro (LV022) encoding the murine sodium iodide symporter (mNIS) cDNA under the spleen focus-forming virus (SFFV) promoter and the puromycin resistance gene (Puro) under the mouse phosphoglycerate kinase (PGK) promoter.

The lentiviral vector used is a self-inactivating (SIN) vector 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).

Mycoplasma Testing:

The CT26.WT-iRFP-Neo/mNIS-Puro cell line has been tested for mycoplasma contamination and is certified mycoplasma free.

Cell Line Authentication:

The parental CT26.WT 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 iRFP and NIS expressing cell line suitable for use as a positive control cell line in fluorescence and iodine uptake assays to verify iRFP or NIS expression in your lentiviral transduced cells.

In vivo: CT26.WT cells form tumors post implantation in mice. The in vivo growth of these tumors can be monitored using noninvasive optical imaging for iRFP or noninvasive, high-resolution 3D PET or SPECT imaging for NIS.

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

NIS Function Assay (Iodine Uptake): Cells were incubated with I-125 for 1h in the presence or absence of KClO4, an inhibitor of NIS mediated iodine uptake. Radioiodine concentrated within the cells was measured with a gamma counter.

Flow Cytometry for iRFP: CT26.WT-iRFP-Neo/mNIS-Puro cells (red) or control (CT26.WT-Fluc-Neo; grey) cells were fixed with paraformaldehyde and analyzed by flow cytometry (20,000 events).