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KC-5816

Ba/F3-mCRBN-humanized-KI-Plus Cell Line

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Datasheet
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Home » Ba/F3-mCRBN-humanized-KI-Plus Cell Line

Background of Ba/F3-mCRBN-humanized-KI-Plus Cell Line

The Ba/F3-mCRBN-humanized-KI cell line is generated by introducing specific humanizing point mutations into the endogenous mouse Crbn locus. This model is designed to support targeted protein degradation research, particularly the high-throughput screening of molecular glue degraders and the investigation of their mechanisms of action. By humanizing CRBN, the cell line recapitulates human-specific drug responses within a murine cellular context. A key advantage of this system is the combination of human-like pharmacological activity with the experimental versatility of Ba/F3 cells. The Ba/F3 background offers ease of genetic manipulation and robust proliferation, facilitating large-scale screening assays. Meanwhile, humanized CRBN expression ensures clinically relevant readouts, improving the translational predictive value of preclinical findings in targeted degradation therapy development.

Specifications

Catalog NumberKC-5816
Cell Line NameBa/F3-mCRBN-humanized-KI-Plus Cell Line
NCBI/UniProt Accession Number58799/Q8C7D2
Clone Number2A4
Host Cell LineBa/F3
DescriptionIn Ba/F3 cells, the endogenous mouse CRBN gene was replaced with the human CRBN coding sequence under the control of an exogenous promoter.
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+8 ng/mL mouse IL-3
Selection MarkerNA
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

Cell Line Generation

Ba/F3-mCRBN-humanized-KI-Plus cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of Ba/F3-mCRBN-humanized-KI cell line stable clone using PCR sequencing.

Figure 2: Characterization of Ba/F3-mCRBN-humanized-KI cell line stable clone using western blot.

Cell Resuscitation

  1. Prewarm culture medium (RPMI1640+10% FBS+8 ng/mL mouse IL-3) 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. Naruse C, Ishibashi O, Asano M, et al. Differential substrate degradation by super-degron: EGFP in wild-type mouse cells, PD-1 requires CRBN humanization. iScience. 2025; 28(7): 112992. doi: 10.1016/j.isci.2025.112992.
  2. Li SR, Fu J, Wang H, et al. IMiD compounds affect CD34+ cell fate and maturation via CRBN-induced IKZF1 degradation. Blood Adv. 2018;2(5):492–504. doi:10.1182/bloodadvances.2017010348. PMID:29606568; PMCID:PMC5876033.

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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