KC-6339

HT1080-FAP-mcherry Cell Line

63962

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Background of HT1080-FAP-mcherry Cell Line

FAP, also known as FAPalpha, is a type II transmembrane glycoprotein that belongs to the serine protease family, which is selectively expressed in reactive stromal fibroblasts of epithelial cancers, granulation tissue of healing wounds, and malignant cells of bone and soft tissue sarcomas. FAP is thought to be involved in the control of fibroblast growth or epithelial-mesenchymal interactions during development, tissue repair, and epithelial carcinogenesis. Acts as a tumor suppressor in melanocytic cells through regulation of cell proliferation and survival in a serine protease activity-independent manner.

Specifications

Catalog Number	KC-6339
Cell Line Name	HT1080-FAP-mcherry Cell Line
NCBI/UniProt Accession Number	NP_004451.2
Clone Number	7#
Host Cell Line	HT1080
Description	Stable HT1080 cell line expressing exogenous FAP-mcherry 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% basal medium+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
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

HT1080-FAP-mcherry cell line was generated using a lentiviral vector expressing the FAP-mcherry sequence.

Characterization

Figure 1: Characterization of FAP overexpression in the HT1080-FAP-mcherry stable clone using FACS.

Figure 2: Characterization of mcherry overexpression in the HT1080-FAP-mcherry stable clone using FACS.

Figure 3: Characterization of FAP mcherry in the HT1080-FAP-mcherry stable clone using PCR sequencing.

Cell Resuscitation

Pre-warm complete culture medium (basal medium and 10% FBS) 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.Puré E, Blomberg R (August 2018). Pro-tumorigenic roles of fibroblast activation protein in cancer: back to the basics. Oncogene. 37 (32): 4343–4357.
2. Xin L, Gao J, Zheng Z, Chen Y, Lv S, Zhao Z, Yu C, Yang X, Zhang R. Fibroblast Activation Protein-α as a Target in the Bench-to-Bedside Diagnosis and Treatment of Tumors: A Narrative Review. Front Oncol. 2021 Aug 19;11:648187.
3. Busek P, Mateu R, Zubal M, Kotackova L, Sedo A (June 2018). Targeting fibroblast activation protein in cancer - Prospects and caveats. Frontiers in Bioscience. 23: 1933–1968.
4. Niedermeyer J, Garin-Chesa P, Kriz M, Hilberg F, Mueller E, Bamberger U, Rettig WJ, Schnapp A (April 2001). Expression of the fibroblast activation protein during mouse embryo development. The International Journal of Developmental Biology. 45 (2): 445–7. PMID 11330865.

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.