Immune Cell Preparation

Preparation of highly active and pure immune cells is a crucial element in cell therapy research and treatment. With extensive experience in immune cell preparation, Cyagen offers a range of services for the preparation of CAR-T, CAR-NK, and universal CAR-T cells, ensuring high quality and efficiency in the development of cell therapies.

Service Offerings:

• T and CAR-T cell preparation
• NK and CAR-NK cell preparation
• Universal CAR-T cell preparation
• Mouse CAR-T cell preparation

Service Advantages:

• Cutting-edge CAR-T/NK cell preparation process producing cells with high positivity rates and excellent functional characteristics
• Advanced UCAR-T cell preparation process generating highly pure and active cells with exceptional positivity rates
• Rigorous quality control standards guaranteeing the reliability and reproducibility of all our immune cell preparation services

T/CAR-T Cell Preparation

T cells are one of the most important adaptive immune cells in the body and play a critical role in fighting against diseases. Chimeric antigen receptor T cell (CAR-T) therapy has made significant breakthroughs in the field of cancer treatment. With extensive experience in T cell preparation, Cyagen can provide high-purity T cells through various methods, which can obtain different proportions of CD4 and CD8 T cell subtypes. In addition, Cyagen offers customized CAR-T cell preparation services, ensuring the anti-tumor function of the CAR-T cells. The CAR-T cell preparation process is as follows:

Deliverables：T and CAR-T cells, experiment report.

Case Study: CD19 CAR-T Cell Preparation

Figure 1. Phenotypic analysis of T cells expanded by two different methods.

Figure 2. Positivity rate detection of CD19 CAR-T cells.

Case Study: GPC3 CAR-T Cell Preparation

Figure 3. GPC3 CAR-T cell positivity rate detection.

UCAR-T Cell Preparation

Currently available CAR-T cell therapies are individualized treatment products with high production costs, long production cycles, high risk of production failure, high prices, and difficulty achieving industrialization. Therefore, research on universal CAR-T cell therapy has gained increasing attention in recent years.

One obstacle to the use of T cells for allogeneic therapy is the serious graft-versus-host disease (GvHD) that can occur after infusion. Current studies have shown that the TCR receptor complex is an important molecule that mediates the occurrence of GvHD, while studies have also shown that T cells in which TCR complex expression is blocked can be used for allogeneic therapy without causing toxic side effects such as GvHD.

Based on CRISPR-Pro technology, Cyagen has established effective TCRα and CD52 knockout processes and, combined with lentivirus transduction technology, can prepare universal CAR-T cells with high purity and high CAR molecule expression to meet the needs of research on different target antigens.

The UCAR-T cell preparation process is as follows:

Deliverables：UT cells and UCAR-T cells, and experiment reports.

Case Study: GPC3 Universal CAR-T Cell Preparation

Figure 4. Efficiency of TCRα and CD52 gene knockout in T cells.

Figure 5. Efficiency of TCRα and CD52 knockout cell enrichment.

Figure 6. Positivity rate of GPC3 UCAR-T cells.

NK/CAR-NK Cell Preparation

NK cells are an important component of innate immunity, playing critical roles in anti-viral infections, anti-tumor responses, and immune regulation. As innate immune cells, NK cells can exert immune functions without prior activation, and often play important roles before adaptive immune cells are activated. Furthermore, the anti-tumor activity of NK cells is not restricted by MHC molecules, making them a promising tool for allogeneic immunotherapy, with safety demonstrated in multiple clinical trials.

Cyagen has established high-purity and high-activity NK cell and CAR-NK cell preparation methods to meet customer demands for NK cell immunotherapy and ADCC studies on antibody drugs.

The CAR-NK cell preparation process is as follows:

Deliverables：NK and CAR-NK cells, and experiment reports.

Case Study: GPC3 CAR-NK Cell Preparation

Figure 7. Statistical analysis of NK cell amplification using two different methods.

Figure 8. Purity detection of NK cells amplified by method 1.

Figure 9. Detection of GPC3 CAR-NK cell positivity rate.

Immortalized CAR Cell Preparation

Large-scale screening of CAR molecules is a labor-intensive and time-consuming process. To reduce the workload, preliminary screening can be carried out using immortalized cells such as Jurkat cells, NK-92 cells or THP-1 cells.

Deliverables：Stable transfected cell lines and an experiment report.

Client-provided information：Target information, scFv antibody sequence (optional), CAR molecule structure information, desired vector information, and required number of cells.

Case Study: Construction and Functional Evaluation of GPC3 CAR-Jurkat Cells

Figure 10. Experimental results of the construction and functional validation of GPC3 CAR-Jurkat cells.

A. Detection of GPC3 CAR-Jurkat cell positivity rate, demonstrating the successful construction of GPC3 CAR-Jurkat cells;

B. Detection of GPC3 antigen expression level on liver cancer cells;

C. Detection of activation marker CD69 and exhaustion markers PD-1 and TIM-3 expression in GPC3 CAR-Jurkat cells co-cultured with liver cancer HepG2 cells, showing that GPC3 CAR-Jurkat cells can be effectively activated by target cells;

D. Results of the analysis in C.