Nalm6-Fluc-Neo/eGFP-Puro is a clonal population of the B cell precursor leukemia cell line Nalm6 that stably expresses firefly luciferase and enhanced green fluorescent protein (eGFP).
The Nalm6-Fluc-Neo/eGFP-Puro cell line was generated by transducing parental Nalm6 cells with two separate lentiviral vectors. The first encoding firefly luciferase and the neomycin resistance gene (LV-Luc2-P2A-Neo (LV011)) and the second encoding enhanced green fluorescent protein and the puromycin resistance gene (LV-eGFP-PGK-Puro (LV031)). Transduced cells were selected using puromycin and G418 followed by clonal selection using a methylcellulose based semi-solid medium.
This cell line has been tested for mycoplasma and no mycoplasma has been detected. The parental Nalm6 cell line was authenticated by short tandem repeat (STR) profiling.
SCID beige mice were injected intravenously with 1×106 Nalm6-Fluc-Neo/eGFP-Puro cells. Bioluminescent and fluorescent imaging was performed at the indicated times using a Xenogen IVIS Spectrum.
Low and high density cell photos (200x).
The indicated number of cells were placed in wells of a 96-well plate. After the addition of 3 mg/mL d-luciferin, the plate was immediately imaged using a Xenogen IVIS Spectrum. The total flux (photons/sec) was plotted as a function of cell number.
Nalm6-Fluc-Neo/eGFP-Puro (green) or isotype control (Nalm6-Fluc-Puro grey) cells were fixed with paraformaldehyde and analyzed by flow cytometry.
Nalm6-Fluc-Neo/eGFP-Puro cells were stained with an anti-HuCD19 (A), anti-HuBCMA (B), anti-HuCD20 (C), anti-HuCD38 (D), or anti-HuGPRC5D (E) antibody (green) or isotype control antibody (grey) and analyzed by flow cytometry.
Complete Growth Medium: ATCC-formulated RPMI-1640 supplemented with 10% fetal bovine serum (FBS), and 1X Penicillin/Streptomycin.
The addition of 1 µg/mL puromycin and 1 mg/mL G418 to the complete growth medium is recommended if maintaining the cells in culture for more than two weeks.
Grow cells in the indicated medium, subculturing as needed to maintain a density between 3×105 and 3×106 cells/mL. Passage cells by dilution in fresh medium with regular complete media changes.
These cells are suitable for in vitro and in vivo experimentation.
The luciferase transgene facilitates noninvasive bioluminescent imaging of implanted cells in vivo and cell-based assays in vitro. eGFP is not recommended for whole animal in-live imaging. Rather, samples can be collected post mortem for analysis by conventional fluorescence microscopy or flow cytometry.
Nalm6 cells form tumors and metastases at multiple sites (liver, lymph nodes, spleen, bone marrow) upon implantation into immunocompromised mice.
These cells can be amplified to generate additional frozen stocks and cryopreservation of low passage stocks in a designated cryopreservation medium is recommended.
These cells were generated by transduction with a self-inactivating Risk Group 3 (RG3) lentiviral vector. All work should be performed under appropriate biosafety conditions per institutional guidelines.
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