CELL LINES
Contributor Information
- Name T.R. Santhosh Kumar
- Institute Rajiv Gandhi Centre for Biotechnology
- Primary citation Lupitha SS, et.al. PMID: 32109534.
Tool Details
- Tool name: U-251/EGFP_LC3
- Tool type: Cell Lines
- Parental cell line: U-251MG
- Organism: Human
- Tissue: Brain
- Gender: Male
- Disease: Cancer
- Morphology: Epithelial
- Growth properties: Adherent
- Model: Transgenic
- CRISPR: No
- Conditional: No
- Application: Tested for LC3B punctae formation by widefield fluorescence microscopy and confocal microscopy. Ideal for real time imaging and high-throughput screening.
- Description: MAP1LC3B (Microtubule associated protein 1 Light Chain 3 ?) is a protein involved in the autophagic process. Under normal physiological conditions the protein is cytosolic and during autophagy it undergoes lipidation to involve in autophagosome elongation. This process can be visualized as LC3B punctae formation. The cell line stably express MAP1LC3B tagged with EGFP and allow easy identification and quantitation of cells with autophagy response using a wide variety of fluorescent imaging platforms including high content imaging. The cell line is an ideal cell tool for autophagy and apoptosis research and support long time imaging of autophagy in live cells.
- Research area: Cancer; Neurodegenerative diseases; Metabolic disorders
- Production details: The product is a human breast adenocarcinoma cell line U-251, transfected to stably express the protein MAP1LC3B tagged with EGFP. Single cell clones with homogeneous expression of the fusion protein were developed thorough colony selection and flow sorting. The EGFP-tag enables easy identification and affinity purification of the recombinant protein. The cell line can be utilized for studying the autophagy process and to screen of potential candidate drugs that induce autophagy.
- For Research Use Only
Target Details
Application Details
- Application: Tested for LC3B punctae formation by widefield fluorescence microscopy and confocal microscopy. Ideal for real time imaging and high-throughput screening.
Handling
- Growth medium: DMEM with 2mM L-Glutamine and 10% FBS (Heat inactivated);
- Temperature: 37C
- Atmosphere: 5% CO2
- Storage medium: Growth medium + 10% DMSO
- Storage conditions: Liquid Nitrogen
- Shipping conditions: Dry Ice
- Initial handling information: Resuscitation: Rapidly thaw the frozen ampoule in a water bath at 37Ă°C for 1-2 minutes. Transfer the contents to a centrifuge tube and slowly add 5-10ml of pre-warmed growth media. Remove a sample for counting. Centrifuge at 100-150g for 2-3 minutes to pellet cells (a centrifugation step to remove the cryoprotectant is essential. Seed at the recommend density, e.g. greater than 10,000 cells per cm2.
- Subculture routine: Subculture non-confluent cultures using trypsin/EDTA and 1:3 to 1:4, i.e. seeding at 1-4 x10,000 cells/cmĂÂ .
- Cultured in antibiotics?: No
Documentation
- Available on request
References
- • Lupitha SS, et.al. PMID: 32109534.
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