Comparative flow cytometry immunophenotyping of ASCs expanded in conventional flasks versus automated bioreactors

Area of Science:

• Regenerative Medicine
• Stem Cell Biology
• Biotechnology

Background:

• Adipose-derived stem cells (ASCs) are vital for regenerative medicine, requiring large-scale in vitro expansion.
• Traditional tissue culture polystyrene (TCP) flasks have limitations for mass ASC production.
• Hollow fiber bioreactors (HFBs) offer an automated, closed system for cell expansion.

Purpose of the Study:

• To compare immunophenotypes of ASCs expanded in HFB systems versus traditional TCP flasks.
• To evaluate the impact of expansion methods on ASC surface marker expression and heterogeneity.
• To assess the suitability of HFB for clinical-grade ASC manufacturing.

Main Methods:

• ASCs from three donors were cultured in both HFB and TCP systems with equivalent population doublings.
• Cell counts compared growth rates between the two systems.
• Multicolor flow cytometry analyzed 15 surface markers and co-expression of specific epitopes.

Main Results:

• Both HFB and TCP systems demonstrated comparable ASC growth rates.
• Most surface marker profiles were similar, except for CD201 downregulation in HFB (p=0.008).
• HFB supported greater immunophenotypical heterogeneity, preserving CD274+ and CD274- subpopulations, unlike TCP which favored CD201+ and CD274+ clones.

Conclusions:

• Expansion methods significantly influence ASC phenotypic composition.
• HFB systems are a promising alternative for large-scale ASC manufacturing due to maintained subpopulation diversity.
• Further functional validation of ASC subtypes and careful selection of expansion platforms are essential for clinical applications.