KC-2789

Ba/F3-EGFR-B2M-HLA-A1101-Cell-Line

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Home » Ba/F3-EGFR-B2M-HLA-A1101-Cell-Line

Background of Ba/F3-EGFR-B2M-HLA-A1101-Cell-Line

B2M, also known as β2 microglobulin, is a component of MHC class I molecules and expressed on nearly all nucleated cell. B2M noncovalently lies beside α3 heavy chain of MHC class I molecules on the cell surface without transmembrane region. HLA-A belongs to the HLA class I heavy chain paralogues. HLA-A is a heterodimer consisting of a heavy chain and a light chai. The heavy chain is anchored in the membrane. HLA-A plays a central role in the immune system by presenting peptides derived from the endoplasmic reticulum lumen so that they can be recognized by cytotoxic T cells. They are expressed in nearly all cells. HLA-A is a Protein Coding gene. Diseases associated with HLA-A include Severe Cutaneous Adverse Reaction and Birdshot Chorioretinopathy. Among its related pathways are SARS-CoV-2 Infection and Infectious disease. 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 NumberKC-2789
Cell Line NameBa/F3-EGFR-B2M-HLA-A1101-Cell-Line
Host Cell LineBa/F3-EGFR
DescriptionStable Ba/F3-EGFR clone expressing exogenous B2M-HLA-A1101 gene
QuantityOne vial of frozen cells (≥2-106/vial)
StabilityStable in culture over a minimum of 10 passages
ApplicationDrug screening and biological assays
Freezing Medium70% RPMI1640+20% FBS+10% DMSO
Propagation MediumRPMI1640+10%FBS+1000µg/mL Hygromycin B
Selection MarkerHygromycin B
MorphologyMostly single, round (some polymorph) cells in suspension
SubcultureSplit saturated culture 1:10 every 3 days; seed out at about 1-3 × 105 cells/mL
Incubation37 °C with 5% CO2
StorageLiquid nitrogen immediately upon receiving
Doubling TimeApproximately 20 hours
Mycoplasma StatusNegative
In Vivo ValidationNA

Cell Line Generation

Ba/F3 EGFR-B2M-HLA-A1101 cell line was generated using a retrovirus vector expressing the human B2M-HLA-A1101 fusion sequence.

Characterization

Figure 1: Characterization of B2M-HLA-A1101 overexpression in the Ba/F3-EGFR stable clone by using FACS.

Cell Resuscitation

  1. Prewarm culture medium (RPMI1640+10%FBS+1000µg/mL Hygromycin B) 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% 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. Wang H, Liu B, Wei J. Beta2-microglobulin(B2M) in cancer immunotherapies: Biological function, resistance and remedy. Cancer Lett. 2021 Oct 1;517:96-104. doi: 10.1016/j.canlet.2021.06.008. Epub 2021 Jun 12. PMID: 34129878.
  2. Manca MA, Simula ER, Cossu D, Solinas T, Madonia M, Cusano R, Sechi LA. Association of HLA-A*11:01, -A*24:02, and -B*18:01 with Prostate Cancer Risk: A Case-Control Study. Int J Mol Sci. 2023 Oct 20;24(20):15398. doi: 10.3390/ijms242015398. PMID: 37895076; PMCID: PMC10607162.

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.
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