MACSductin™ Reagent

Sensitive: transduction of target cells with low titer virus preparations
Efficient: magnetic capture of virus particles enables maximal concentration at target cells resulting in efficient transduction
Specific: MACS® Technology ensures specific transduction of target cells

Overview

MACSductin™ Reagent consists of polycationic, magnetic beads that help to efficiently transduce primary cells and cell lines using adeno- or retro-/lentiviral vectors. It enables transduction of target cells with low-titer virus preparations. The efficiency is superior compared to standard transduction methods.

Details

Background information
During infection of target cells with native or recombinant viruses in cell culture, the major factor limiting infection is the diffusion of the virus particle to the cell surface. MACSductin™ Reagent in combination with MACS® Technology enables close colocalization of virus and target cells within MACS Columns to ensure optimal transduction efficiencies. The polycationic, magnetic MACSductin Reagent binds to the adeno- or retro-/lentivirus. The target cells are specifically labeled with antibody-conjugated, superparamagnetic MACS MicroBeads. As both target cells and virus are retained by the magnetic field on the column in close proximity to each other, the virus can quickly attach to the cell surface resulting in a high infection or transduction efficiency.

Detailed procedure
Virus aliquots are incubated with MACSductin™ Reagent. Magnetically labeled target cells are then added to the virus-MACSductin Reagent complex and the mixture is loaded onto an MS or LS Column placed in a magnetic field. The column is then washed to remove potential contaminants i.e. from unpurified virus-containing cell culture supernatant and removed from the magnetic field. Finally, the virus-cell complexes are eluted in fresh cell culture medium and cultured before proceeding to downstream applications. The basic principle is illustrated in figure 1.

Downstream applications
Cells transduced with MACSductin Reagent can be cultured under standard cell culture conditions. They can be further analyzed and characterized using standard molecular biology applications. Genetically modified cells can also be introduced into rodent animal models.

Columns

MS Columns or LS Columns

Figure 1

Principle of transduction with MACSductin Reagent and MACS Technology

Figure 2

Transduction of primary human CD34⁺ cells with unpurified cell culture supernatant containing lentiviral vector
Primary CD34⁺ cells were isolated from PBMCs using the CD34 MicroBead Kit, human. Cells were cultured for 24 hours in 24-well plates at 1×10⁵ cells per mL in medium supplemented with Flt-3L, SCF, IL-3, and IL-6. 1×10⁵ cells, magnetically labeled with CD34 MicroBeads, were then transduced in MS Columns with unpurified supernatant containing a GFP-encoding, replication-incompetent SIN lentiviral vector at an MOI of 50, complexed with different amounts of MACSductin Reagent. GFP expression of transduced CD34⁺ cells was measured 48 hours post-transduction in a MACSQuant® Analyzer.

Figure 3

Transduction of human cell lines with purified adenoviral vector
1×10⁶ HuT78 and A549 cells were magnetically labeled with CD30 and CD44 MicroBeads, respectively. In parallel, purified adenoviral vector particles encoding GFP were complexed with MACSductin Reagent. The A549 cells were then transduced in MS Columns at a pMOI (physical particles per cell) of 50 while HuT78 cells were transduced at a pMOI of 500. GFP expression of transduced cells was increased up to 30-fold 24 hours post-transduction as measured by flow cytometry (data courtesy of Dr. Florian Kreppel and Dr. Sigrid Espenlaub, Department of Gene Therapy, University of Ulm, Germany).