KC-4319

MC38-TSHR Cell Line

44262

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Background of MC38-TSHR Cell Line

TSHR (Thyroid Stimulating Hormone Receptor) is a Protein Coding gene. The protein encoded by this gene is a membrane protein and a major controller of thyroid cell metabolism. Thyroid-stimulating hormone (TSH), through activation of its G-protein-coupled thyrotropin receptor (TSHR), controls the synthesis of thyroid hormone—an essential metabolic hormone. Diseases associated with TSHR include Hypothyroidism, Congenital, Nongoitrous, 1 and Hyperthyroidism, Familial Gestational.

Specifications

Catalog Number	KC-4319
Cell Line Name	MC38-TSHR Cell Line
NCBI/UniProt Accession Number	NM_000369.5
Clone Number	2#
Host Cell Line	Mouse MC38 cell line
Description	Stable MC38 clone expressing exogenous human TSHR 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% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS + 5μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
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

MC38-TSHR cell line was generated using a lentiviral vector expressing the human TSHR sequence.

Characterization

Figure 1: Characterization of human TSHR overexpression in MC38-TSHR stable clones using FACS.

Figure 2: Characterization of TSHR in the MC38-TSHR stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (DMEM supplemented with 10% FBS and 5μg/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.Duan, J., Xu, P., Luan, X. et al. Hormone- and antibody-mediated activation of the thyrotropin receptor. Nature 609, 854–859 (2022).
2. Smits G, Govaerts C, Nubourgh I, Pardo L, Vassart G, Costagliola S. Lysine 183 and glutamic acid 157 of the TSH receptor: two interacting residues with a key role in determining specificity toward TSH and human CG. Mol Endocrinol. 2002 Apr;16(4):722-35. doi: 10.1210/mend.16.4.0815. PMID: 11923469.
3. Tuncel M. Thyroid Stimulating Hormone Receptor. Mol Imaging Radionucl Ther. 2017 Feb 9;26(Suppl 1):87-91. doi: 10.4274/2017.26.suppl.10. PMID: 28117293; PMCID: PMC5283706.