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Updated: Mar 2, 2026
Technique for Obtaining Mesenchymal Stem Cell from Adipose Tissue and Stromal Vascular Fraction Characterization in Long-Term Cryopreservation
Published on: December 30, 2021
Las enzimas DisoNNex y TrypLE son intercambiables como agentes de recolección para células madre mesenquimales
Ellen Mønsted Johansen1, Stine Bangsgaard1, Laura Lykke Lethager1
1Cardiology Stem Cell Centre University Hospital of Copenhagen, Copenhagen, Denmark.
Background Aims:
Cell therapies based on adipose tissue-derived mesenchymal stromal cells (ASCs) are promising candidates for the treatment of numerous diseases. Relevant to the potential of these therapies is the development of consistent and efficient ASC-manufacturing processes, with enzymatic cell dissociation being a critical step. DisoNNex, a novel cell dissociation enzyme based mainly on chymotrypsin activity, offers a Good Manufacturing Practice (GMP)-compliant alternative to existing dissociation methods; its impact on ASC characteristics remains yet to be explored.
Methods:
The performance of DisoNNex was compared with the current dissociation industry standard, TrypLE Select Enzyme 1x (TrypLE; Gibco/Thermo Fisher Scientific, Waltham, MA, USA). The DisoNNex concentration matching the dissociation kinetics of TrypLE was defined using time-lapse microscopy and image analysis. ASCs from five donors were harvested using either TrypLE or DisoNNex across two passages. Assays assessed immunophenotype, apoptosis, secretion of vascular endothelial factor, indoleamine 2,3-dioxygenase expression and suppression of lymphocyte proliferation. Multi-omics (bulk RNA sequencing, proteomics, metabolomics and lipidomics) were included in the study.
Results:
DisoNNex at 15 µg/mL matched TrypLE in dissociation kinetics and maintained ASC viability. No significant differences were observed in functional assays. Multi-omics analysis revealed only four differentially expressed genes of 12509 protein-coding genes, and no significant differences in proteins, metabolites or lipids were found. The Multi-Omics Factor Analysis plot showed no clustering according to the dissociation enzymes used for harvest.
Conclusions:
DisoNNex is a relevant alternative to TrypLE for ASC harvest, resulting in equivalent yields and cell quality under the tested conditions. These findings support the potential of DisoNNex in GMP-compliant manufacturing of ASC-based cell therapies and provide starting conditions for its implementation in ASC processes. Nonetheless, full comparability studies within an established production process are pending.
