Cat.No. : CSC-RT0101
Host Cell: HEK293 Target Gene: SLC1A5
Size: 1x10^6 cells/vial, 1mL Validation: Sequencing
| Cat. No. | CSC-RT0101 |
| Cell Line Information | This cell line is a stable cell line with a homozygous knockout of human SLC1A5 using CRISPR/Cas9. |
| Target Gene | SLC1A5 |
| Gene ID | 6510 |
| Genotype | SLC1A5 (-/-) |
| Host Cell | HEK293 |
| Cell Type | Epithelial |
| Size | 1x10^6 cells/vial, 1mL |
| Sequencing Result | Homozygous: 34 bp deletion in exon |
| Species | Homo sapiens (Human) |
| Revival | Rapidly thaw cells in a 37°C water bath. Transfer contents into a tube containing pre-warmed media. Centrifuge cells and seed into a 25 cm2 flask containing pre-warmed media. |
| Media Type | Cells were cultured in DMEM supplemented with 10% fetal bovine serum. |
| Growth Properties | Cells are cultured as a monolayer at 37°C in a humidified atmosphere with 5% CO2. Split at 80-90% confluence, approximately 1:3-1:6. |
| Freeze Medium | Complete medium supplemented with 10% (v/v) DMSO |
| Mycoplasma | Negative |
| Format | One frozen vial containing millions of cells |
| Storage | Liquid nitrogen |
| Safety Considerations |
The following safety precautions should be observed. 1. Use pipette aids to prevent ingestion and keep aerosols down to a minimum. 2. No eating, drinking or smoking while handling the stable line. 3. Wash hands after handling the stable line and before leaving the lab. 4. Decontaminate work surface with disinfectant or 70% ethanol before and after working with stable cells. 5. All waste should be considered hazardous. 6. Dispose of all liquid waste after each experiment and treat with bleach. |
| Ship | Dry ice |
Alanine-serine-cysteine transporter 2 (ASCT2, SLC1A5) is the major transporter of glutamine in cancer cells. In this study, researchers developed a novel monoclonal antibody (mAb) that recognizes the extracellular domain of human ASCT2 and investigated whether ASCT2 could be a therapeutic target for KRAS mutant cancers. Rats were immunized with RH7777 rat hepatoma cells expressing human ASCT2 fused to green fluorescent protein (GFP). Splenocytes from immunized rats were fused with P3X63Ag8.653 mouse myeloma cells, and hybridoma cells secreting Ab3-8 mAb were established for screening and cloning. The mAb reacted with RH7777 transfectants expressing ASCT2-GFP protein in a GFP intensity-dependent manner. Ab3-8 reacted with various human cancer cells but not with noncancerous mammary epithelial cells or ASCT2 knockout HEK293 and SW1116 cells. In SW1116 and HCT116 human colon cancer cells with KRAS mutations, treatment with Ab3-8 reduced intracellular glutamine transport, phosphorylation of AKT and ERK, and inhibited tumor growth of these cells in athymic mice. Ab3-8 inhibition of in vivo tumor growth was not observed in HT29 colon and HeLa uterine cancer cells with wild-type KRAS. These results suggest that ASCT2 is an excellent therapeutic target for KRAS mutant cancers.
Ab3-8 did not react with ASCT2 (SLC1A5) Knockout Cell Line-HEK293 (ASCT2-KO-HEK293) and SW1116 cells (Figure 1E). In addition, Ab3-8 reacted with various human cancer cells but was weakly reactive with non-cancerous cells (Figure 1F). Among these cell lines, SW1116, HCT116, DLD-1, and HCT15 cells contained KRAS mutations, while the other cell lines had wild-type KRAS.
Figure 1. E, ASCT2-KO HEK293 and SW1116 cells were reacted with Ab3-8 and subsequently with PE-conjugated anti-rat IgG. The reactivity of Ab3-8 with these cells was analyzed by flow cytometry. F, FCM analysis of cell surface expression of ASCT2 protein in various human cell lines. (Hara, Yuta, et al. 2020)
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