KC-5739

MC38-HLA-A0201 Cell Line

59015

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Background of MC38-HLA-A0201 Cell Line

HLA-A (Major Histocompatibility Complex, Class I, A) is a Protein Coding gene. HLA-A belongs to the HLA class I heavy chain paralogues. This class I molecule is a heterodimer consisting of a heavy chain and a light chain (beta-2 microglobulin). The heavy chain is anchored in the membrane. Class I molecules play 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. Diseases associated with HLA-A include Severe Cutaneous Adverse Reaction and Hla Modifier.

Specifications

Catalog Number	KC-5739
Cell Line Name	MC38-HLA-A0201 Cell Line
Clone Number	15#
Host Cell Line	Mouse MC38 Cell Line
Description	Stable MC38 clone expressing exogenous human HLA-A0201 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 + 100μg/mL Hygromycin B
Selection Marker	Hygromycin B
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 20 hours
Mycoplasma Status	Negative

Cell Line Generation

MC38-HLA-A0201 cell line was generated using a lentiviral vector expressing the human HLA-A0201 sequence.

Characterization

Figure 1: Characterization of human HLA-A0201 overexpression in the MC38 HLA A0201 stable clone using FACS.

Figure 2: Characterization of human HLA A0201 in the MC38 stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (DMEM supplemented with 10% FBS and 100μg/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 1:4-1:8 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.Möller P, Hämmerling GJ. The role of surface HLA-A,B,C molecules in tumour immunity. Cancer Surv. 1992;13:101-27. PMID: 1423320.
2. Mashayekhi P, Omrani MD, Yassin Z, Dehghanifard A, Ashouri L, Aghabozorg Afjeh SS, Shabanzadeh Z. Influence of HLA-A, -B, -DR Polymorphisms on the Severity of COVID-19: A Case-Control Study in the Iranian Population. Arch Iran Med. 2023 May 1;26(5):261-266. doi: 10.34172/aim.2023.40. PMID: 38301089; PMCID: PMC10685865.
3. Palmer WH, Norman PJ. The impact of HLA polymorphism on herpesvirus infection and disease. Immunogenetics. 2023 Jun;75(3):231-247. doi: 10.1007/s00251-022-01288-z. Epub 2023 Jan 3. PMID: 36595060; PMCID: PMC10205880.