KC-6003

KP4-GFP-Luc2 Cell Line

60263

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Background of KP4-GFP-Luc2 Cell Line

Green fluorescent protein (GFP) is a protein that exhibits bright green fluorescence when exposed to light in the blue to ultraviolet range. The label GFP traditionally refers to the protein first isolated from the jellyfish Aequorea victoria and is sometimes called avGFP. Green fluorescence protein (GFP) is a protein composed with 238 amino acids and isolated from Aequorea victoria. GFP emits green fluorescent spontaneously.
Luciferase is an oxidative enzyme that can produce bioluminescence with addition of luciferin, but don’t need an external light source unlike of fluorescent proteins. Photo emission can be detected directly by light sensitive device. such as luminometer or modified microscope. Luciferase is widely used in many fields of biological research, such as transcriptional activity, kinase or other enzyme activity, cellular ATP level, whole animal imaging.

Specifications

Catalog Number	KC-6003
Cell Line Name	KP4-GFP-Luc2 Cell Line
Clone Number	2#
Host Cell Line	KP4
Description	Stable KP4 clone expressing exogenous GFP and Luciferase gene.
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10%FBS
Selection Marker	NA
Morphology	Epithelial
Subculture	Split saturated culture 1:2~1:3 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative

Cell Line Generation

KP4-GFP-Luc2 cell Line was generated using retrovirus vector expressing GFP and Luciferase sequence.

Characterization

Figure 1: Characterization of KP4-GFP-Luc2 Cell Line stable clone using FACS.

Figure 2: Characterization of KP4-GFP-Luc2 Cell Lines stable clone using Bright-Lite Luciferase Assay System in the conditions of different cell number.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 supplemented with 10% FBS)in a 37°C water bath.
2. Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
3. Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
4. Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
5. Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
6. Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
7. Incubate the flask at 37°C, 5% CO₂ incubator.
8. Split saturated culture 1:2~1:3 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI1640 + 20% FBS + 10% DMSO) fresh immediately before use.
2. Keep the freezing medium on ice and label cryovials.
3. Transfer cells to a sterile, conical centrifuge tube, and count the cells.
4. Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
5. Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
6. Aliquot 1 mL of the cell suspension into each cryovial.
7. Freeze cells in the Cool Cell freezing container overnight in a -80°C freezer.
8. Transfer vials to liquid nitrogen for long-term storage.

References

1.Salih A (2019). "Fluorescent Proteins". In Cox G (ed.). Fundamentals of Fluorescence Imaging. Boca Raton: Jenny Stanford Publishing. p. 122. doi:10.1201/9781351129404. ISBN 9781351129404. S2CID 213688192..
2.Fan Y, Li Y, Yao X, Jin J, Scott A, Liu B, Wang S, Huo L, Wang Y, Wang R, Pool Pizzi M, Ma L, Shao S, Sewastjanow-Silva M, Waters R, Chatterjee D, Liu B, Shanbhag N, Peng G, Calin GA, Mazur PK, Hanash SM, Ishizawa J, Hirata Y, Nagano O, Wang Z, Wang L, Xian W, McKeon F, Ajani JA, Song S. Epithelial SOX9 drives progression and metastases of gastric adenocarcinoma by promoting immunosuppressive tumour microenvironment. Gut. 2023 Apr;72(4):624-637. doi: 10.1136/gutjnl-2021-326581. Epub 2022 Aug 24. PMID: 36002248.