KC-2779

Calu6 B7H3 KO 3A3 Cell Line

27167

Home » Calu6 B7H3 KO 3A3 Cell Line

Background of Calu6 B7H3 KO 3A3 Cell Line

B7-H3 is a 316 amino acid-long type I transmembrane protein, existing in two isoforms determined by its extracellular domain. In mice, the extracellular domain consists of a single pair of immunoglobulin variable (IgV)-like and immunoglobulin constant (IgC)-like domains. B7-H3 mRNA is expressed in most normal tissues. In contrast, B7-H3 protein has a very limited expression on normal tissues because of its post-transcriptional regulation by microRNAs. However, B7-H3 protein is expressed at high frequency on many different cancer types (60% of all cancers).

Specifications

Catalog Number	KC-2779
Cell Line Name	Calu6 B7H3 KO 3A3 Cell Line
Host Cell Line	Calu6
Description	Stable Calu6 clone with human B7H3 gene knockout, No.3A3
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% MEM + 0.01Mm NEAA+20% FBS + 10% DMSO
Propagation Medium	MEM + 0.01Mm NEAA + 10% FBS
Selection Marker	N/A
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:5 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 48 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

Calu-6 -B7H3 knockout cell line was generated using a CRISPR method.

Characterization

Figure 1: Characterization of B7H3 knockout in Calu-6 using PCR sequencing.

Figure 2: Characterization of B7H3 knockout in Calu-6 using FACS.

Cell Resuscitation

Prewarm culture medium (MEM + 0.01Mm NEAA + 10% FBS)in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:4-1:5 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% MEM + 0.01Mm NEAA+20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage

References

Zhang GB, Chen YJ, Shi Q, Ma HB, Ge Y, Wang Q, Jiang Z, Xu Y, Zhang XG (June 2004). Human recombinant B7-H3 expressed in E. coli enhances T lymphocyte proliferation and IL-10 secretion in vitro. Acta Biochimica et Biophysica Sinica. 36 (6): 430–6. doi:10.1093/abbs/36.6.430. PMID 15188059.

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.