KC-4036

Ba/F3-GHR-Cell-Line

40530

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Background of Ba/F3-GHR-Cell-Line

GHR, also known as GHBP, this gene encodes a member of the type I cytokine receptor family, which is a transmembrane receptor for growth hormone. Binding of growth hormone to the receptor leads to receptor dimerization and the activation of an intra- and intercellular signal transduction pathway leading to growth. Mutations in this gene have been associated with Laron syndrome, also known as the growth hormone insensitivity syndrome (GHIS), a disorder characterized by short stature. In humans and rabbits, but not rodents, growth hormone binding protein (GHBP) is generated by proteolytic cleavage of the extracellular ligand-binding domain from the mature growth hormone receptor protein. Multiple alternatively spliced transcript variants have been found for this gene. Research has shown the role of the growth hormone receptor (GHR) gene in skeletal malocclusion and its significant influence on the growth of the maxilla and the mandible in both sagittal and vertical dimensions.
Ba/F3 cell, a murine interleukin-3 dependent pro-B cell line, is a popular system for exploring both kinases and their inhibitors, because some protein kinases can render the Ba/F3 cells to be depended on the activation of the kinases instead of IL-3 supplement, while their inhibitors can antagonize the kinase-dependent growth effects.

Specifications

Catalog Number	KC-4036
Cell Line Name	Ba/F3-GHR-Cell-Line
Clone Number	2-4#
Host Cell Line	Ba/F3
Description	Stable Ba/F3 clone expressing exogenous GHR 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	RPMI 1640 + 10% FBS + 8ng/mL mouse IL-3 + 1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Mostly single, round (some polymorph) cells in suspension
Subculture	Split saturated culture 1:10 every 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 20 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

Ba/F3-GHR-cell-Line was generated using retrovirus vector expressing human GHR sequence.

Characterization

Figure 1: Characterization of GHR overexpression in the Ba/F3 human GHR stable clone using FACS.

Figure 2: Characterization of GHR in the Ba/F3-GHR stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (RPMI 1640 supplemented with 10% FBS and 8ng/mL mouse IL-3 and 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 1:10 every 3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI-1640 + 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. Unterberger CJ, McIlwain SJ, Tsourkas PK, Maklakova VI, Prince JL, Onesti A, Hu R, Kopchick JJ, Swanson SM, Marker PC. Conditional gene regulation models demonstrate a pro-proliferative role for growth hormone receptor in prostate cancer. Prostate. 2023 Apr;83(5):416-429.
2. Perez Garrido N, Pujana M, Berger M, Ramírez P, Guercio G, Belgorosky A, Marino R. Growth hormone receptor gene polymorphism. Spontaneous catch up growth in small for gestational age patients. Medicina (B Aires). 2021;81(4):574-580. English.
3. George AM, Felicita AS, Priyadharsini VJ, P A, Tr PA. Role of the Growth Hormone Receptor (GHR) Gene in Skeletal Class II Malocclusion and Its Significant Influence on the Skeletal Facial Profile in Both the Sagittal and Vertical Dimensions: A Systematic Review. Cureus. 2024 Feb 5;16(2):e53596.