Increased 3T3 Cell Growth Following

Electromagnetic Stimulation with Mitoplicator™

A single, 3-­‐minute stimulation with ultrashort, pulsed electrical fields proved to accelerate the growth rate of the 3T3.L1 cell line. This inductive, non-­‐contact technology leads to larger cell batches within a shorter time frame.

INTRODUCTION

The Mitoplicator™ instrument (figure 1) promotes cell proliferation by means of nanosecond pulsed electrical fields. There is no need for direct contact with the cell culture, because cells are simulated within a standard culture vial.

Here we describe the result following single stimulation of the adherent cell line 3T3.L1 cells (ATCC -­‐ CL-­‐173).

RESULTS

Pre-­‐confluent trypsinized cells, contained in closed standard culture vials (Cryotubes) were exposed to pulsed electrical fields for 3 min., seeded at 5000 cells/ml and subsequently cultured for 3 days.

In figure 2 can be seen that control cells duplicated twice, whereas most stimulated cells completed 3 duplications over 3 days. This leads to a 1,75-­‐fold higher yield in cell number in treated cultures versus controls.

There was no effect on cell viability (trypan blue dye exclusion) and no morphological changes were observed using the light microscope.

CONCLUSION

A single pulsed electrical field stimulation is an effective way to increase the yield of cell cultures in standard culture conditions or in a shorter time frame.

                           Figure 2.
3T3.L1 cell proliferation (n = 3) following a 3 min. stimulation

METHODS

Cells. 3T3.L1 (ATCC CRL-­‐173) were cultured following ATCC guidelines 1. (DMEM: Gibco 41965-­‐039; FBS: Gibco10500-­‐064).

Stimulation.Pre-­‐confluent cell monolayers were trypsinized, blocked and suspended. 2 ml aliquots were dispensed in cryovials, closed and just prior to placement inside the Mitoplicator™, resuspended by turning all the vials twice. The control vial was placed aside the instrument for the duration of the stimulation.

Seeding. Of both treated and control samples, 5 000 cells were seeded side-­‐by-­‐side and in triplicate in 24-­‐well plates for manual counting.

Manual counting. Monolayers were trypsinized, blocked and suspended. Trypan blue solution was added. Suspensions were dispensed into a glass Burker-­‐type chamber. Viable (trypan blue negative) and dead (trypan blue positive) cells were hand counted in quadruplicate.

ABOUT MITOPLICATOR™ & ECPR TECHNOLOGY

The Enhanced Cell Proliferation Reactor (ECPR) technology was discovered, validated & developed by the Vabrema founders since 2009 and in close collaboration with the Eindhoven Technical University (TU/e).

Today, ECPR technology is available for R&D purposes as the Mitoplicator™ instrument and for fermenter-­‐scale cell or protein production (assay/engineering project).

Proof of principles for ECPR technology includes mammalian & human cell lines, primary cells & isolates, stem cells, plant seedlings, algae. Vabrema actively pursues an R&D plan, aiming at widening ECPR applicability.

Please inquire with us if ECPR technology could be to the benefit of your R&D or company value chain.