KC-5907

293T-mem-KLK2 Cell Line

59771

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Background of 293T-mem-KLK2 Cell Line

Human kallikrein-related peptidase 2 (KLK2) is a tryptic serine protease predominantly expressed in prostatic tissue and secreted into prostatic fluid, a major component of seminal fluid, it is a Protein Coding gene. This gene encodes a member of the grandular kallikrein protein family. Kallikreins are a subgroup of serine proteases that are clustered on chromosome 19. Members of this family are involved in a diverse array of biological functions. The protein encoded by this gene is a highly active trypsin-like serine protease that selectively cleaves at arginine residues. This protein is primarily expressed in prostatic tissue and is responsible for cleaving pro-prostate-specific antigen into its enzymatically active form. Diseases associated with KLK2 include Prostate Disease and Prostate Cancer.

Specifications

Catalog Number	KC-5907
Cell Line Name	293T-mem-KLK2 Cell Line
NCBI/UniProt Accession Number	NM_005551.5
Clone Number	2#
Host Cell Line	293T
Description	Stable 293T cell line expressing exogenous KLK2 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% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS + 1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial-like
Subculture	Split saturated culture 1:4-1:8 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
In Vivo Validation	NA

Cell Line Generation

293T-mem-KLK2 cell line was generated using a lentiviral vector expressing the GPI-anchored KLK2 sequence.

Characterization

Figure 1: Characterization of mem KLK2 overexpression in the 293T-mem-KLK2 stable clone using FACS(JNJ78278343-KLK2-hIgG1, kyinno, Cat#KB-1777).

Figure 2: Characterization of mem KLK2 in the 293T stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (DMEM + 10% FBS + 1μg/mL Puromycin)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 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% DMEM + 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 CoolCell freezing container overnight in a -80°C freezer.
8. Transfer vials to liquid nitrogen for long-term storage.

References

1. Lövgren J, Valtonen-André C, Marsal K, Lilja H, Lundwall A. Measurement of prostate-specific antigen and human glandular kallikrein 2 in different body fluids. J Androl. 1999 May-Jun;20(3):348-55. PMID: 10386814.
2. Batra J, O'Mara T, Patnala R, Lose F, Clements JA. Genetic polymorphisms in the human tissue kallikrein (KLK) locus and their implication in various malignant and non-malignant diseases. Biol Chem. 2012 Dec;393(12):1365-90. doi: 10.1515/hsz-2012-0211. PMID: 23667899.
3. Skala W, Utzschneider DT, Magdolen V, Debela M, Guo S, Craik CS, Brandstetter H, Goettig P. Structure-function analyses of human kallikrein-related peptidase 2 establish the 99-loop as master regulator of activity. J Biol Chem. 2014 Dec 5;289(49):34267-83. doi: 10.1074/jbc.M114.598201. Epub 2014 Oct 16. PMID: 25326387; PMCID: PMC4256358.

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.