Species: Human 

Cell type: Melanoma

Transgenes: Human sodium iodide symporter (hNIS) with neomycin resistance (Neo) for selection with G418 and enhanced green fluorescent protein (eGFP) with puromycin resistance (puro) for selection with puromycin

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

 Description: Mel624-CMVhNIS-Neo/eGFP-Puro is a polyclonal population of the human melanoma Mel624 cell line transduced with a lentiviral vector encoding the human sodium iodide symporter (hNIS) cDNA under the cytomegalovirus (CMV) promoter linked to the neomycin resistance gene via an IRES and LV-eGFP-PGK-Puro (LV031) encoding enhanced green fluorescent protein (eGFP) under the spleen focus-forming virus (SFFV) promoter and puromycin resistance (Puro) under the 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).

Recommended uses:

In vitro: This is a high hNIS/eGFP expressing clone suitable for use as a positive control cell line in I-125 uptake assays to validate NIS expression and fluorescence assays to validate GFP expression in your lentiviral transduced cells.

In vivo: This human melanoma cell line grows as subcutaneous xenografts in aythmic nude mice and its growth can be monitored noninvasively using high-resolution 3D SPECT or PET imaging. 

References on NIS imaging:

1. Fruthwirth et al. A whole body dual modality radionuclide optical strategy for preclinical imaging of metastasis and heterogeneous treatment responses in different. microenvironments. J. Nucl. Med 2014. 55(4): 686-94.

2. Penheiter et al. The sodium iodide symporter (NIS) as an imaging reporter for gene, viral and cell-based therapies. Curr Gene Ther. 2012, 12(1):33-47.

 

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)

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

 

Flow Cytometry for eGFP: Mel624-CMVhNIS-Neo/eGFP-Puro (green) or control (grey) cells were fixed with paraformaldehyde and analyzed by flow cytometry (20,000 events).