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Improved expansion of T cells in culture when isolated with an equipment-free, high-throughput, flow-through microfluidic module versus traditional density gradient centrifugation

Published:January 16, 2019DOI:https://doi.org/10.1016/j.jcyt.2018.12.004

      Highlights

      • Flow-through device recovers more lymphocytes than density gradient centrifugation.
      • Cell recoveries were 85% CD3+, 89% CD19+ and 97% CD56+ with this new approach.
      • Microfluidic-isolated cells grew faster than gradient-isolated cells in culture.
      • Overall, the new method gives 2X higher cumulative cell yield after 7-day culture.
      • Devices can process 200 mL of sample in less than 1 h, with no pumping mechanism needed.

      Abstract

      Background

      The isolation of lymphocytes – and removal of platelets (PLTs) and red blood cells (RBCs) – from an initial blood sample prior to culture is a key enabling step for effective manufacture of cellular therapies. Unfortunately, currently available methods suffer from various drawbacks, including low cell recovery, need for complex equipment, potential loss of sterility and/or high materials/labor cost.

      Methods

      A newly developed system for selectively concentrating leukocytes within precisely designed, but readily fabricated, microchannels was compared with conventional density gradient centrifugation with respect to: (i) ability to recover lymphocytes while removing PLTs/RBCs and (ii) growth rate and overall cell yield once expanded in culture.

      Results

      In the optimal embodiment of the new microfluidic approach, recoveries of CD3+, CD19+ and CD56+ cells (85%, 89% and 97%, respectively) were significantly higher than for paired samples processed via gradient-based separation (51%, 53% and 40%). Although the removal of residual PLTs and RBCs was lower using the new approach, its enriched T-cell fraction nevertheless grew at a significantly higher rate than the gradient-isolated cells, with approximately twice the cumulative cell yield observed after 7 days of culture.

      Discussion

      The standardization of each step of cellular therapy manufacturing would enable an accelerated translation of research breakthroughs into widely available clinical treatments. The high-throughput approach described in this study – requiring no ancillary pumping mechanism nor expensive disposables to operate – may be a viable candidate to standardize and streamline the initial isolation of lymphocytes for culture while also potentially shortening the time required for their expansion into a therapeutic dose.

      Graphical Abstract

      Key Words

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