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HCT116-Fluc-Neo/eGFP-Puro

Species: Human

Cell type: Colorectal carcinoma

Transgenes: Firefly luciferase (Fluc) with neomycin resistance (Neo) for selection with G418 and enhanced green fluorescent protein (eGFP) with puromycin resistance for selection with puromycin

Media: DMEM, 10% FBS, 1% Pen/Strep, 0.5 mg/mL G418, 1 μg/mL puromycin

Description: HCT116-Fluc-Neo/eGFP-Puro is a polyclonal population of the human colorectal carcinoma cell line HCT116 (ATCC® CCL-247™) transduced with LV-Fluc-P2A-Neo (LV011) encoding firefly luciferase (Fluc) cDNA under the spleen focus-forming virus (SFFV) promoter linked to the neomycin resistance gene (Neo) via a P2A cleavage peptide and LV-eGFP-PGK-Puro (LV031) encoding enhanced green fluorescent protein (eGFP) cDNA under the SFFV promoter and the puromycin resistance gene (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).

Mycoplasma Testing: This cell line has been tested for mycoplasma contamination and is certified mycoplasma free.

Cell Line Authentication: Authentication of the parental HCT116 cell line was performed by short tandem repeat (STR) profiling with 9 STR loci including CSF1PO, D13S317, D16S539, D5S818, D7S820, TH01, TPOX, vWA and sex chromosome marker Amelogenin. STR profiling of HCT116 cells are verified and there is no interspecies cross contamination detected.

Recommended uses:

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

In vivo: This human colorectal cell line grows as xenografts in immunocompromised mice. Growth of these tumors can be monitored in vivo using noninvasive bioluminescent imaging with D-luciferin substrate or optical imaging for GFP fluorescence. Note: In life optical imaging 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)

Flow cytometry: Histograms showing GFP expression in HCT116-Fluc-Neo/eGFP-Puro (green) versus control HCT-Fluc-Neo cells (gray)

Luciferase Assay: 104, 105, or 106 cells were placed in wells of a 96-well plate and 0.3 mg of d-luciferin was added to the indicated wells. The plate was immediately imaged using a Xenogen IVIS Spectrum.

Bioluminescent images of a subcutaneous HCT116-Fluc-Puro tumor in an athymic mouse

107 HCT116-Fluc-Puro cells (Imanis Life catalog #CL008) were injected subcutaneously into the right flanks of female NCR athymic mice. Mice were imaged at day 0 on the day of cell implantation and at days 7 and 16 using a Perkin Elmer IVIS® Spectrum system, at 10-15 minutes post intraperitoneal injection of D-luciferin at 150 mg/kg. Tumor size was measured using calipers. Data from a representative mouse (ID#4772) is shown.

 

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With quality reagents, services, and support, we have you covered at all stages of your study. Our expert scientists can help you select the best reagents for your needs and offer tips to optimize your experiments and achieve superior results. Our wide-range of reporter gene and oncolytic virus reagents undergo rigorous quality control testing so you can be confident in the quality of the products you receive. And with our competitive prices, you can complete your study for less.

  • “Our group has, and continues to, use NIS as a noninvasive reporter for cell transplantation studies in mice and in pigs. Imanis has provided expert technical and analytical support for this research, and has allowed us to publish our research in high impact journals, including Science Translational Medicine.” – Dr. Raymond Hickey, Mayo Clinic

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Use Fewer Animals

Our Reduction Campaign

At Imanis, we are committed to promoting the practice of the 3Rs in animal research. Learn how we are decreasing the use of animals and research as well as saving up to 15% on your orders. Continue reading...

What people say about Imanis

Our group has, and continues to, use NIS as a noninvasive reporter for cell transplantation studies in mice and in pigs. Imanis has provided expert technical and analytical support for this research, and has allowed us to publish our research in high impact journals, including Science Translational Medicine..

– Dr. Raymond Hickey, Mayo Clinic

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