Clinical-grade T cell gene-engineering via electroporation and transduction

CliniMACS Prodigy T Cell Engineering (TCE) allows for complex gene modifications by combining electroporation and transduction. The automated and closed process covers the entire workflow from cell isolation, activation, electroporation and/or viral transduction to cell expansion and harvest.

The CliniMACS Electroporator expands the cell processing capabilities of the CliniMACS Prodigy Platform. It is an add-on to the CliniMACS Prodigy and enables large-scale transfection in an automated and closed fashion. The electroporation parameters can be flexibly adapted to optimize transfection protocols for different gene-engineering technologies, e.g., TALENs, CRISPR/Cas, Zinc finger, Sleeping Beauty, siRNA, and more. Additional viral cell transduction is an option for further and more complex gene engineering approaches. To guarantee high quality cell manufacturing we provide all ancillary materials for the entire process. The combination of the CliniMACS Prodigy and the CliniMACS Electroporator brings GMP-compliant gene-engineering to the next level. 

Four easy steps to your engineered T cell product

Workflow overview of the T cell engineering (TCE) manufacturing process on the CliniMACS Prodigy and CliniMACS Electroporator

The CliniMACS Prodigy Platform enables the entire workflow for T cell engineering, from cell separation and activation, to genetic modification and cell expansion. All cell processing steps are automated, ensuring a highly reproducible and standardized manufacturing process.

Cell separation on the CliniMACS Prodigy is a simple and easy method to magnetically isolate T cells with a high purity. Advantages of this completely automated method include: 

  • Efficient separation of CD4+ and CD8+ cells without inducing activation
  • Removal of contaminating immune cells to enhance T cell engineering efficiency
  • No magnetic bead removal necessary

MACS® GMP T Cell TransAct™ is a colloidal polymeric nanomatrix conjugated to humanized, recombinant CD3 and CD28 agonists. It is a ready-to-use reagent that has been designed to activate and expand enriched T cell populations or resting human T cells from peripheral blood mononuclear cells (PBMCs). Thanks to its nanoscale structure the reagent can be sterile filtered and excess reagent can conveniently be removed by washing the TransAct activated cells.

MACS GMP T Cell TransAct is directly applicable for the use in automated culture systems, such as the CliniMACS Prodigy.

The CliniMACS Prodigy TCE integrates electroporation into automated cell manufacturing. At the same time, it offers the freedom to perform viral transduction on the target cell or to combine both methods for more complex genetic modification approaches.    

Electroporation

Non-viral gene engineering opens new possibilities for genetically modified cell-based drug products. One very common method of non-viral gene engineering is electroporation of cells of interest.

The genetic information is transferred into the cell via an applied electrical charge which induces pores in the cell membrane. The CliniMACS Prodigy TCE facilitates an optimized transfection performance by enabling flexible adaptation of different electroporation parameters while maintaining a high cell viability.

Graphical depiction of example pulses that can be performed with the CliniMACS Electroporator. 

The pulse parameters such as voltage, length, mode, polarity, and burst length can be configured by the user to be optimized for the chosen gene-editing substrate (e.g. CRISPR/Cas, TALENs, mRNA, etc.).

For automated electroporation single and double pulses can be selected via the CliniMACS Prodigy touch screen. The pulses themselves can be defined either as square wave or or burst pulses with variable lengths and voltages. For the first pulse the user is free to additionally choose the polarity of the pulse.  

Table: Flexible pulse parameters are available to optimize the electroporation efficiency.  

 1st Pulse2nd Pulse
Voltage [V]50-100050-500
Length [µs]5-100,0000-100,000
ModeSquare and burstSquare and burst
PolarityUnipolar and bipolar-
Burst length [µs]5-100,0005-100,000

Viral transduction

To achieve long-term gene expression through stable genomic insertion, two types of gene transduction tools are commonly used: gamma-retroviral vectors (RV) or lentiviral vectors (LV). CliniMACS Prodigy TCE allows for the combination of electroporation and viral transduction with each of these vectors.  

The transduction enhancer Vectofusin®-1 is the perfect choice to enhance your RV transduction efficiency. It is compatible with both static and spinoculation protocols.

Cell expansion takes place in the cell cultivation chamber, which is part of the CliniMACS Prodigy Tubing Set in order to maintain a closed system. As part of the TCE activity matrix, the cell culture duration as well as the medium supply and exchange are flexibly programmable.

Ideal cultivation and expansion of engineered T cells rely on the strong synergy of MACS GMP T Cell TransAct™, TexMACS™ GMP Medium, and MACS GMP Cytokines.

TexMACS GMP Medium

  • Specifically developed for T cell cultivation
  • Serum- and xeno-component free
  • Pharmaceutical-grade human serum albumin
  • QC functionality test on every batch

MACS GMP Cytokines

  • Lot-to-lot consistency and lot-specific certificates of analysis
  • Designed according to the recommendations of <USP 1043> on ancillary materials
  • Manufactured and tested under a certified ISO 13485 quality system

At the end of the CliniMACS Prodigy T Cell Engineering, the cells are harvested in a solution of your choice. 

Express Modes of the MACSQuant Instruments allow for fully automated and standardized flow cytometry processes.

In-process and quality controls (IPC/QC) are required for consistent cell manufacturing. Integrated sampling pouches on the CliniMACS Tubing Sets allow for samples to be collected at any time during and at the end of the cell manufacturing process. Miltenyi Biotec provides complete solutions with its MACS Flow Cytometers, a wide portfolio of MACS IVD Antibodies, a time-saving and standardized staining procedure with StainExpress antibody cocktails, and automated flow cytometry analysis with Express Modes for clinical flow cytometry. 

When using a MACSQuant® Instrument, Express Modes allow for fully automated and standardized flow cytometry processes. The MACSQuant Instrument and the MACS Flow Cytometry Portfolio are for research use only. MACS IVD Antibodies are produced according to the quality management system ISO 13485.

Disclaimer

For the manufacture and use of gene-engineered T cells, national and international legislation and regulations must be followed. The manufacturer has to validate the QC method. Miltenyi Biotec as the provider of products for the T cell engineering does not give any recommendation regarding the use of the manufactured cells for therapeutic purposes and does not make any claim regarding a clinical benefit.

Alzubi, J. et al. (2020) Automated generation of gene-edited CAR T cells at clinical scale. Mol. Ther. Methods Clin. Dev. 20: 379–388.

Schwarze, L. I. et al. (2021) Automated production of CCR5-negative CD4+ T cells in a GMP-compatible, clinical-scale for treatment of HIV-positive patients. Gene Ther.

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.

Seems like you are coming from USA!
Do you want to visit our website in your country?