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Protein Sciences

Uniting Drug Development Expertise with Antibody & Protein Production Services

T Cell Mate Service

High-Throughput, High-Titer CHO Expression of T Cell-Related Proteins

 

T cell receptor (TCR) therapy, a novel approach for targeting intracellular proteins via major histocompatibility complex (MHC), is gaining traction in the drug development field. Traditionally, E. coli was used for TCR expression, but encountered challenges such as low yield and limited throughput.

 

To overcome these challenges, we have developed the T Cell Mate service. Utilizing our high-titer CHO expression system, this service enables the high-throughput production of soluble TCRs, TCR fusion proteins and single-chain trimers (SCT) with high yield. Additionally, the T Cell Mate service offers refolded peptide-MHC complexes (RF-pMHC) production in E. coli.

T-Cell Reception

Key Features of T Cell Mate Service:

  • High-titer CHO production of soluble TCR proteins, TCR fusion proteins, and SCTs
  • High-throughput and high yield
  • Protein delivery within 3-4 weeks

The T-Cell Mate service offers high-titer CHO expression of soluble TCR proteins, TCR fusion proteins, and SCTs.

T Cell Mate Service Details:

Service Item Deliverables Duration QC Request A Quote
sTCR, TCR-Ab fusions Purified proteins ranging from mg to grams per request 3-4 weeks Bradford, SDS-PAGE, SEC-HPLC, Endotoxin, LC-MS (optional) Request A Quote
SCT Purified proteins ranging from mg to grams per request 3-4 weeks Bradford, SDS-PAGE, SEC-HPLC, Endotoxin, LC-MS
RF-pMHC (E. Coli) Purified proteins ranging from mg to grams per request 3-4 weeks Bradford, SDS-PAGE, LC-MS

Case Study #1: High-Titer Expression of Soluble TCR in CHO Cells

 

In this case study, we demonstrated the high-titer expression of sTCR proteins in CHO cells using our T Cell Mate service.

Figure A: The first gel shows one-arm Fc fusion protein with a yield of 1,092 mg/L after single-step purification. The second gel shows His-tagged TCR with a yield of 111.2 mg/L following affinity purification. The third gel shows single-step, affinity-purified scFv infused with TCR with a yield of 90.7 mg/L.

T-Cell Mate service demonstrated high-titer expression of soluble TCR and TCR fusion proteins in CHO cells.

Case Study #2: High-Purity, High-Yield Production of Mouse SCTs in CHO Cells

 

In this case study, we demonstrated the production of a mouse single-chain-trimer (SCT) with high purity and yield, obtained from a 20 mL CHO culture. It was achieved through 2-step purification using our T Cell Mate service.

Figure B: After the initial purification using a Ni column, we achieved a purity of 82.42% with a yield of 18.3 mg. Following the second SEC purification step, the purity was increased to 99.57%, yielding 10 mg of SCT. The purity was confirmed by NR SDS-PAGE.

T-Cell Mate service demonstrated high-purity and high-yield production of a mouse single-chain-trimer (SCT) proteins in CHO cells.

Frequently Asked Questions for T Cell Mate Service

Q: Is Fc component involved in zipper design for MHC II protein production?

A: While Fc could technically be used, we recommend using the Leucine Zipper. This method involves using a very small peptide which is easily removed once the protein has been produced.

Q: Does your platform provide other membrane proteins besides TCR?

A: Our service is focused on producing soluble TCRs without the transmembrane domain. These TCRs are suitable for reagent or therapeutic purposes. However, we’re open to evaluating membrane protein request.

Q: Do the TCRs have an engineered disulfide bond in the constant domains?

A: Yes, our TCRs feature an engineered disulfide bond in the constant domains. Additionally, we remove the native disulfide bond present in the stalk region.

Q: How do you compare the activities of secreted MHC vs. refolded MHC?

A: Generally, the consensus is that these single-chain trimers (SCTs) perform comparably to refolded peptide-MHC complexes. Despite this, the production of refolded MHC complexes in E. coli remains the standard method. However, for HTP MHC production, CHO cells are still the preferred system, offering a cost-effective and rapid solution.

Q: How do you ensure that the peptide remains bound to the MHC during refolding?

A: To verify, we employ LC-MS analysis to ensure the complex integrity, and we do not store the complex in a buffer with excess peptides.

Q: What considerations are important when selecting an expression system for TCRs, such as E. coli versus mammalian systems?

A: To produce TCRs, we prefer using CHO expression system due to its ability to produce TCRs that are naturally folded and glycosylated with high titers. This approach is generally more effective than attempting to refold TCRs produced in E. coli.

Q: Without the need for post-translational modifications, is there still a reason to use mammalian expression systems?

A: Yes, the CHO system is preferred for producing naturally folded TCRs and supports high-throughput TCR production, making it ideal for early-stage research and screening purposes.

Q: Can you produce various TCR types?

A: Yes, our system is fully capable of expressing various TCR types, such as murine TCRs and hybrid constructs.

Q: How do you ensure the purity of the TCRs?

A: To ensure the purity of TCRs, we employ a variety of analytical methods, such as SEC-HPLC, SEC-MALS, SDS-PAGE, and Intact Mass analysis. These methods provide a comprehensive assessment of the TCR purity, ensuring premium-quality products.

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