Takara Lenti-X™ Transduction Sponge, 24 Rxns

The Lenti-X Transduction Sponge is a macroporous alginate structure that facilitates rapid and efficient lentiviral transduction without spinoculation or the use of a chemical transduction enhancer. It encourages the co-localization of target cells and lentivirus, eliminating biotransport issues associated with standard lentiviral transduction methods. A simple, easy-to-use protocol reduces cell handling and requires smaller total reaction volumes while producing transduction efficiencies that are comparable to or improved over traditional methods.

Simple transduction workflow using the Lenti-X Transduction Sponge.Cells and virus are mixed prior to application to the sponge. The transduction mixture is incubated for 1 hr, followed by the addition of media and overnight incubation for 16–24 hr. No spinoculation is required. The next day, healthy, transduced cells are released using an optimized Release Buffer that depolymerizes the alginate matrix. Cells are then ready for subsequent analysis or continued culture.

The Lenti-X Transduction Sponge comes in a convenient format for use with a 24-well tissue culture plate. Panel A.Each kit comprises 24 individual sponges, each capable of facilitating transduction of up to 1 x 107cells.Panel B. Every sponge features a complex microfluidic pore structure with pore sizes ranging from 20–300 µm. The image shown is at 150X magnification.Panel C. Simple placement of the transduction sponge into a well before applying the sample.Panel D.The transduction mix, consisting of 1 x 105–1 x 107cells and virus, is added to the sponge and incubated for 1 hr before the addition of media and incubation for 16–24 hr.Panel E. Cell release is facilitated by transferring the sponge to a 15 mL conical tube and adding Release Buffer, followed by a short wash step.

The Lenti-X Transduction Sponge facilitates transduction at equivalent levels to spinoculation in multiple cell types. Panel A.1 x 106Jurkat cells were transduced with lentivirus expressing ZsGreen1 at the indicated multiplicities of infection (MOIs) using spinoculation or the Lenti-X Transduction Sponge. The spinoculation cultures were centrifuged for 90 min at 1400xgat 32°C in the presence of 8 μg/mL polybrene. All transductions were analyzed at 48 hr post-transduction for ZsGreen1 expression by FACS.Panel B. To further compare sponge transduction to spinoculation ("Spin" in the figure), various cell lines were transduced with ZsGreen1 lentivirus at several MOIs, as described in Panel A.

The Lenti-X Transduction Sponge enhances primary T-cell transduction efficiency while preserving viability. Panel A.Human primary T cells were activated using MHC tetramer, anti-CD3/CD28-coated beads, Retronectin reagent (RN) + CD3, or CD3 only. Activation levels indicated by percentage CD69+ expression at 48 hr are displayed. Then, 1 x 106activated cells were transduced with lentivirus encoding ZsGreen1 at the specified multiplicity of infection (MOI) using the transduction sponge for 24 hr. FACS analysis was performed at 48 hr post-transduction to determine %ZsGreen1 expression.Panel B.Viability at 48 hr post-transduction as determined by 7-AAD staining and FACS analysis.

The Lenti-X Transduction Sponge demonstrates consistent transduction efficiency across varying cell amounts and transduction times with low inter-sponge variability. Panel A. Jurkat cells were transduced with a lentivirus encoding ZsGreen1 at specified cell numbers and multiplicities of infection (MOIs) using either a RetroNectin reagent + spinoculation (RN + spin) protocol or the transduction sponge for 24 hr.Panel B. Increasing numbers of Jurkat cells were transduced for 4, 9, 24, or 48 hr using the transduction sponge with a lentivirus expressing ZsGreen1 (MOI=10).Panel C. 1 x 106Jurkat cells were transduced with lentivirus expressing ZsGreen1 (MOI=2.5) in six replicates weekly for 4 weeks to assess inter-run variability (%CV=12.6). For all experiments, ZsGreen1 expression was analyzed by FACS 48 hr post-transduction to determine %ZsGreen positive cells and mean fluorescence intensity (MFI). The average ZsGreen1 expression across all replicates is shown in red (AVG).