Clinical-grade Treg cell manufacturing

  • Clinical-scale Treg cell enrichment and isolation 
  • Convenient combination of our CliniMACS® Platform with MACS® GMP Products
  • Customized strategies  

Regulatory T (Treg) cells are a specialized subpopulation of T cells that act to suppress immune responses, thereby maintaining homeostasis and self-tolerance. Their unique function has made them an interesting target cell population for immunotherapy in a number of different indications, including prevention and treatment of graft-versus-host disease (GvHD) after haploid stem cell transplantation (HSCT)1–4, suppression of solid organ rejection after transplantation5–9, and treatment of autoimmune diseases like type 1 diabetes.

To address the variety of Treg cell applications, Miltenyi Biotec provides a broad range of tools for clinical use. By combining the CliniMACS Cell Manufacturing Platform with MACS GMP Products and our GMP-compliant cell sorter MACSQuant® Tyto®, you can now significantly enhance your clinical-grade Treg cell manufacturing possibilities.

Video

MACS Academy: Improve your clinical-grade Treg manufacturing

Tune into this webinar as we provide you with comprehensive solutions for manufacturing clinical-grade Treg cells. Learn about how you can utilize our CliniMACS Platform and MACS GMP Products for a range of applications. 

The three steps to your final clinical-grade Treg cell product

The clinical-grade Treg cell manufacturing process consists of three steps, in which Treg cell isolation counts as the most challenging. To meet the need of GMP-compliance, automation, scalability, and customizable cell manufacturing, we provide two approaches for isolation of regulatory T cells. Furthermore we offer unique and comprehensive solutions for Treg cell activation, cultivation, and analysis.

1. Clinical-grade Treg separation
 

Approach A: Cell separation using the CliniMACS Plus

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Isolation of polyclonal Treg cells 

Cell separation on the CliniMACS Plus Instrument is an easy and GMP-compliant method to magnetically isolate your CD25high Treg cells, with or without prior depletion of CD8 or CD19 cells with a high purity and recovery.

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Advantages of this method: 

  • Robust large-scale Treg cell enrichment
  • Convenient isolation procedure
  • Maintenance of Treg function and phenotype
  • Pre-depletion of CD8+ (and/or CD19+) cells
  • Efficient enrichment of cells highly expressing CD25 
     

Approach B: Cell separation using the CliniMACS Prodigy and MACSQuant Tyto Cell Sorter

With Treg cells being more and more used as treatment options to suppress solid organ rejection after transplantation, for autoimmune diseases, and as CAR Treg cells, the demand for ultra-pure antigen-specific Tregs has recently increased tremendously. 

Miltenyi Biotec now offers solutions to meet the needs of the field. The combination of the CliniMACS Prodigy for pre-enrichment and the MACSQuant Tyto Cell Sorter for subsequent high-purity cell sorting guarantees full sterility of your medical product.  

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Automated cell separation and sorting 


Isolation of a ultra-pure Treg cell subpopulation using the CliniMACS Prodigy LP-25 Pre-enrichment Process and the MACSQuant Tyto Cell Sorter.

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Advantages of this method: 

  • Robust ultra-pure isolation of Treg cell subpopulation (>98%)
  • Efficient enrichment of CD25+ cells and fluorescent labeling in a closed and automated system using CliniMACS Prodigy LP-25 Pre-Enrichment
  • GMP-compliant cell sorting using the MACSQuant Tyto Cell Sorter
  • Maintenance of Treg cell functionality and phenotype

2. Clinical-grade Treg cell expansion
 

In order to improve the purity and quantity of clinical-grade Treg cells, we offer GMP-compatible expansion products. Ideal cultivation and expansion of Treg cells rely on the strong synergy of:

  • MACS GMP ExpAct Treg Kit
  • TexMACS™ GMP Medium,
  • MACS GMP Cytokines and
  • MACS GMP Rapamycin

MACS GMP ExpAct Treg Kit

The MACS GMP ExpAct Treg Kit has been designed to efficiently expand Treg cells after a CliniMACS CD25 enrichment procedure while maintaining FoxP3 expression. It can be used in combination with TexMACS GMP Medium and MACS GMP Rapamycin, which inhibits the expansion of conventional T cells and thereby enhances the proportion of expanded Treg cells in culture.

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Customer report
Clinical-grade separation and expansion of regulatory T cells for clinical studies on cell therapy in solid organ transplantation

Henrieta Fazekasova, Sarah Thirkell, Katie Lowe, Andrew Bushell, and Giovanna Lombardi, Immunoregulation laboratory, MRC Centre for Transplantation, King’s College London, Guy’s Hospital, London, UK

MACS GMP Rapamycin

Profit from MACS GMP Rapamycin to enhance the proportion of FoxP3+ Treg cells and to inhibit the expansion of conventional T cells during the cultivation and expansion step. We offer 100 nM ready-to-use MACS GMP Rapamycin to improve your culturing.
 

TexMACS GMP Medium

To enhance your Treg cell culturing, we offer TexMACS GMP medium which is specifically developed for T cell cultivation. TexMACS GMP medium is serum- and xeno-component free, but it is supplemented with pharmaceutical-grade human serum albumin. To support you with the consistently high standards, a QC functionality test is done on every batch.
 

MACS GMP Cytokines (IL-2)

Our MACS GMP Cytokines are designed according to the recommendations of <USP 1043> on ancillary materials and manufactured and tested under a certified ISO 13485 quality system. In addition, we provide you with a lot-to-lot consistency and lot-specific certificates of analysis. 
 

3. In-process and quality control 
 

Treg cell suppression assay

The Treg Suppression Inspector has been developed for the in vitro functional characterization of human CD4+ CD25+ regulatory T cells.
 

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Treg cell in vitro suppression assay

  • Optimized T cell stimulation reagent for Treg suppression assays
  • Based on anti-Biotin MACSiBead™ Particles
  • Analysis of Treg cell suppression activity by reduction of responder T cell proliferation

Flow cytometry analysis

Treg Detection Kits  

Our Treg Detection Kits are optimized antibody kits with pre-titrated antibodies that ensure a reliable flow cytometric analysis of Treg cells. The kits are ready-to-use cocktails, which conveniently reduce pipetting steps. 
 

REAfinity™ Recombinant Antibodies 

REAfinity Antibodies are recombinant antibodies that provide superior lot-to-lot consistency and purity compared to mouse or rat hybridoma-derived, monoclonal antibodies. They have been recombinantly engineered to produce highly specific antibodies that require no FcR blocking step. Additionally, they all have the same IgG1 isotype, requiring less isotype controls.
 

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Treg characterization with REAfinity Antibodies

  • Greater reproducibility: High purity and lot-to-lot consistency with less background
  • Easy handling: Eliminates tedious and costly Fc receptor blocking steps
  • One universal isotype: Convenient and cost-saving

References

  1. Martelli, M. F. et al. (2014) HLA‑haploidentical transplantation with regulatory and conventional T‑cell adoptive immunotherapy prevents acute leukemia relapse. Blood 124(4): 638–644
  2. Martelli, M. F. et al. (2015) Next generation HLA-haploidentical HSCT. Bone Marrow Transplant. 50: 63–66.
  3. Theil, A. et al. (2015) Adoptive transfer of allogeneic regulatory T cells into patients with chronic graft‑versus‑host disease. Cytotherapy 17(4): 473–486.
  4. Hannon, M. et al. (2014) Infusion of clinical‑grade enriched regulatory T cells delays experimental xenogeneic graft‑versus‑host disease. Transfusion 54(2): 353–363.
  5. Nadig, S.N. et al. (2010) In vivo prevention of transplant arteriosclerosis by ex vivo-expanded human regulatory T cells. Nat. Med. 16: 809.
  6. Sagoo, P. et al. (2011) Human regulatory T cells with alloantigen specificity are more potent inhibitors of alloimmune skin graft damage than polyclonal regulatory T cells. Sci. Transl. Med. 3: 83ra42.
  7. Issa, F. et al. (2010) Ex vivo expanded human regulatory T cells prevent the rejection of skin allografts in a humanized mouse model. Transplantation 90: 1321.
  8. Wu, D.C. et al. (2013) Ex vivo expanded human regulatory T cells can prolong survival of a human islet allograft in a humanized mouse model. Transplantation 96: 707.
  9. Safina, N. et al. (2016) Successful expansion of functional and stable regulatory T cells for immunotherapy in liver transplantation. Oncotarget 7(7): 7563–7577.
  10. Bluestone J.A. (2015) Type 1 diabetes immunotherapy using polyclonal regulatory T cells. Sci. Transl. Med. 7, 315ra189.
  11. Putnam A. L. et al. (2013) Clinical grade manufacturing of human alloantigen-reactive regulatory T cells for use in transplantation. A. J. Transplant. 13: 3010–3020.
  12. Battaglia, M. et al. (2005) Rapamycin selectively expands CD4+CD25+FoxP3+ regulatory T cells. Blood 105: 4743–4748.

Related information

Unless otherwise specifically indicated, Miltenyi Biotec products and services are for research use only and not for therapeutic or diagnostic use. For complete regulatory notes, please click here.

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