KC-6341

CHOK1-mem-cyno-KLK3 Cell Line

63963

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Background of CHOK1-mem-cyno-KLK3 Cell Line

Kallikrein-related peptidase 3 (KLK3), also known as prostate-specific antigen (PSA), is the most useful biomarker for prostate cancer (PCa). KLK3 (Kallikrein Related Peptidase 3) is a Protein Coding gene. Early studies suggest that KLK3 is able to inhibit angiogenic processes, which is most likely dependent on its proteolytic activity. KLK3 is an important component in human semen. This protein is secreted by the epithelial cells of the prostate and its main function is to promote sperm liquefaction by decomposing seminal vesicle coagulation protein. It plays a crucial role in the reproductive process. The expression of KLK3 is strictly regulated by androgens and is commonly used as a core biomarker for prostate cancer screening in clinical practice. Diseases associated with KLK3 include Prostate Disease and Prostatitis.

Specifications

Catalog Number	KC-6341
Cell Line Name	CHOK1-mem-cyno-KLK3 Cell Line
NCBI/UniProt Accession Number	Q6DT45
Clone Number	2#
Host Cell Line	CHOK1
Description	Stable CHOK1 clone expressing exogenous mem cyno KLK3 gene
Quantity	One vial 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 + 10μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 days
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 24 hours
Mycoplasma Status	Negative

Cell Line Generation

CHOK1-mem-cyno-KLK3 cell line was generated using a lentiviral vector expressing the mem cyno KLK3 sequence.

Characterization

Figure 1: Characterization of mem cyno KLK3 overexpression in the CHOK1-mem-cyno-KLK3 stable clone using QPCR.

Figure 2: Characterization of mem cyno KLK3 in the CHOK1 stable clone using PCR sequencing.

Cell Resuscitation

Pre-warm complete culture medium (RPMI1640 + 10% FBS + 10μg/mL Puromycin) in a 37°C water bath.
Rapidly thaw the cryovial in a 37°C water bath for 1-2 minutes with gentle agitation.
Transfer the vial to a biosafety cabinet, and disinfect the exterior with 70% ethanol.
Aseptically transfer the cell suspension dropwise into a sterile centrifuge tube containing 9.0 mL of pre-warmed complete medium.
Centrifuge at approximately 125 × g for 5–7 minutes at room temperature, carefully aspirate the supernatant without disturbing the cell pellet.
Gently resuspend the pellet in an appropriate volume of complete medium and transfer the suspension into a T25 flask.
Incubate the flask in a 37°C in a humidified 5% CO₂ incubator.
Assess cell viability and morphology after 24 hours. If cells appear healthy, replace the medium with fresh medium supplemented with the appropriate selective antibiotic.
Subculture the cells at a ratio of 1:4-1:8 every 2-3 days upon reaching 80%–90% confluency.

Cell Freezing

Prepare the freezing medium (70% basal medium, 20% FBS and 10% DMSO) freshly before use.
Pre-chill the freezing medium on ice and label the cryovials accordingly.
Transfer the cell suspension to a sterile conical tube and perform a cell count to determine total viability and density.
Centrifuge the cells at 250×g for 5 minutes at room temperature; carefully aspirate the supernatant.
Gently resuspend the cell pellet in chilled freezing medium, ensuring a minimum cell density of 3×10⁶ cells/mL.
Aliquot 1 mL of the cell suspension into each pre-labeled cryovial.
Place the cryovials into a CoolCell® container and store at -80°C overnight for controlled-rate cooling.
Transfer the cryovials to the liquid nitrogen for long-term storage the following day.

References

1. Koistinen H, Künnapuu J, Jeltsch M. KLK3 in the Regulation of Angiogenesis-Tumorigenic or Not? Int J Mol Sci. 2021 Dec 17;22(24):13545. doi: 10.3390/ijms222413545. PMID: 34948344; PMCID: PMC8704207.
2. Jha SK, Rauniyar K, Chronowska E, Mattonet K, Maina EW, Koistinen H, Stenman UH, Alitalo K, Jeltsch M. KLK3/PSA and cathepsin D activate VEGF-C and VEGF-D. Elife. 2019 May 17;8:e44478. doi: 10.7554/eLife.44478. PMID: 31099754; PMCID: PMC6588350.
3. Li H, Fei X, Shen Y, Wu Z. Association of gene polymorphisms of KLK3 and prostate cancer: A meta-analysis. Adv Clin Exp Med. 2020 Aug;29(8):1001-1009. doi: 10.17219/acem/121521. PMID: 32869960.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.