KC-5656

LNCaP-STEAP2 Cell Line

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Background of LNCaP-STEAP2 Cell Line

STEAP2 (STEAP2 Metalloreductase) is a Protein Coding gene. This gene is a member of the STEAP family and encodes a multi-pass membrane protein that localizes to the Golgi complex, the plasma membrane, and the vesicular tubular structures in the cytosol. It is highly over-expressed in various types of cancer and it up-regulation hindered cellular proliferation, invasion and metastasis abilities by inhibiting EMT process and suppressing PI3K/AKT/mTOR signaling pathway. Diseases associated with STEAP2 include Hepatic Adenomas, Familial and Prostate Cancer.

Specifications

Catalog Number	KC-5656
Cell Line Name	LNCaP-STEAP2 Cell Line
Clone Number	2#
Host Cell Line	Human LNCaP cell line
Description	Stable LNCaP cell line expressing exogenous human STEAP2 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 + 2μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Lymphoblast
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 56-84 hours
Mycoplasma Status	Negative

Cell Line Generation

LNCaP-STEAP2 cell line was generated using a lentiviral vector expressing the human STEAP2 sequence.

Characterization

Figure 1: Characterization of human STEAP2 overexpression in the LNCaP stable clone using FACS(Kyinno, Cat#KB-1646).

Figure 2: Characterization of human STEAP2 in the LNCaP STEAP2 stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 2μg/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 1:2-1:3 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 CoolCell freezing container overnight in a -80°C freezer.
8. Transfer vials to liquid nitrogen for long-term storage.

References

1. Torrez CZ, Easley A, Bouamar H, Zheng G, Gu X, Yang J, Chiu YC, Chen Y, Halff GA, Cigarroa FG, Sun LZ. STEAP2 promotes hepatocellular carcinoma progression via increased copper levels and stress-activated MAP kinase activity. Sci Rep. 2024 Jun 3;14(1):12753. doi: 10.1038/s41598-024-63368-2. PMID: 38830975; PMCID: PMC11148201.
2. Yang Q, Ji G, Li J. STEAP2 is down-regulated in breast cancer tissue and suppresses PI3K/AKT signaling and breast cancer cell invasion in vitro and in vivo. Cancer Biol Ther. 2020;21(3):278-291. doi: 10.1080/15384047.2019.1685290. Epub 2019 Nov 7. PMID: 31696760; PMCID: PMC7012168.
3. Liu T, Niu X, Li Y, Xu Z, Chen J, Xu G. Expression and prognostic analyses of the significance of STEAP1 and STEAP2 in lung cancer. World J Surg Oncol. 2022 Mar 28;20(1):96. doi: 10.1186/s12957-022-02566-6. PMID: 35346237; PMCID: PMC8962583.