Culture expansion of adipose derived stromal cells. A closed automated Quantum Cell Expansion System compared with manual flask-based culture.

Mandana Haack-Sørensen,Morten Juhl,Bjarke Follin,Annette Ekblond,Jens Kastrup,Rebekka H Søndergaard,Sonja K Brorsen

doi:10.1186/s12967-016-1080-9

Mandana Haack-Sørensen, Morten Juhl + Show 5 more

Open Access

https://doi.org/10.1186/s12967-016-1080-9

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Abstract

BackgroundAdipose derived stromal cells (ASCs) are a rich and convenient source of cells for clinical regenerative therapeutic approaches. However, applications of ASCs often require cell expansion to reach the needed dose. In this study, cultivation of ASCs from stromal vascular fraction (SVF) over two passages in the automated and functionally closed Quantum Cell Expansion System (Quantum system) is compared with traditional manual cultivation.MethodsStromal vascular fraction was isolated from abdominal fat, suspended in α-MEM supplemented with 10% Fetal Bovine Serum and seeded into either T75 flasks or a Quantum system that had been coated with cryoprecipitate. The cultivation of ASCs from SVF was performed in 3 ways: flask to flask; flask to Quantum system; and Quantum system to Quantum system. In all cases, quality controls were conducted for sterility, mycoplasmas, and endotoxins, in addition to the assessment of cell counts, viability, immunophenotype, and differentiation potential.ResultsThe viability of ASCs passage 0 (P0) and P1 was above 96%, regardless of cultivation in flasks or Quantum system. Expression of surface markers and differentiation potential was consistent with ISCT/IFATS standards for the ASC phenotype. Sterility, mycoplasma, and endotoxin tests were consistently negative. An average of 8.0 × 107 SVF cells loaded into a Quantum system yielded 8.96 × 107 ASCs P0, while 4.5 × 106 SVF cells seeded per T75 flask yielded an average of 2.37 × 106 ASCs—less than the number of SVF cells seeded. ASCs P1 expanded in the Quantum system demonstrated a population doubling (PD) around 2.2 regardless of whether P0 was previously cultured in flasks or Quantum, while ASCs P1 in flasks only reached a PD of 1.0. Conclusion: Manufacturing of ASCs in a Quantum system enhances ASC expansion rate and yield significantly relative to manual processing in T-flasks, while maintaining the purity and quality essential to safe and robust cell production. Notably, the use of the Quantum system entails significantly reduced working hours and thereby costs.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-016-1080-9) contains supplementary material, which is available to authorized users.

Highlights

Adipose derived stromal cells (ASCs) are a rich and convenient source of cells for clinical regenerative therapeutic approaches
Comparison of cell yield in culture flasks and Quantum system Yield of cells obtained during passage 0 (P0) and P1 were compared for three donors
This loss is based on a direct calculation of lines volumes according to the manufacturer; only 70% of loaded cells enter the actual bioreactor for attachment and cell culture

Summary

Introduction

Adipose derived stromal cells (ASCs) are a rich and convenient source of cells for clinical regenerative therapeutic approaches. Applications of ASCs often require cell expansion to reach the needed dose. In vitro expansion of ASCs leads to a multipotent cell population with the capacity to differentiate into several cell lineages. Development of effective clinical cellular therapies using human autologous or allogeneic ASCs requires manufacturing of large numbers of cells. Production of a therapeutically relevant cell product requires in vitro expansion to generate clinically relevant cell numbers. Cell processing must be performed in compliance with Good Manufacturing Practices (GMP) guidelines. This requires GMP-compliant reagents and materials and standardised processes with high reproducibility [6, 7]

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