Optimal Condition of Isolation from an Adipose Tissue-Derived Stromal Vascular Fraction for the Development of Automated Systems

Abstract

BACKGROUND: The stromal vascular fraction (SVF) isolated from adipose tissue, which contains stem cells as well as other cell types, has been applied in various research fields. Although different enzymatic concentrations and treatment durations have been applied to isolate the SVF, optimal conditions have not been established. Thus, we aimed to establish the optimal conditions for isolation of the SVF from adipose tissue by automated systems. METHODS: The SVF was collected from removed adipose tissues of five donors during surgery. The SVF was treated with 0.1% or 0.2% collagenase type I for 20, 40, or 60 min. Then, colony forming unit (CFU) assays and flow cytometry were performed to characterize the adipose stem cells (ASCs). A cytokine array was used to investigate the correlation between colony-formation ability and the secretion of isolated ASCs. RESULTS: Treatment with 0.1% collagenase type I for 60 min resulted in a higher SVF yield, whereas treatment with 0.1% collagenase for 40 min resulted in higher CFU values. In addition, expression of interleukin (IL)-6, IL-8, and monocyte chemoattractant protein-1 in the SVF was higher in the high-CFU group than in the low-CFU group. CONCLUSION: The optimal conditions for isolation of the SVF from adipose tissue were treatment with 0.1% collagenase type I for 40 min. We identified the conditions required for efficient SVF isolation based on high CFU values, and our results will facilitate the development of automated systems.

Development and Validation of a Fully GMP-Compliant Process for Manufacturing Stromal Vascular Fraction: A Cost-Effective Alternative to Automated Methods

“…The best cell viability was achieved by using the highest concentration and the longest incubation time for collagenase. Consistently, as reported by Lee and colleagues, SVF viability was shown to decrease with increasing treatment duration, but this effect is significant only for a duration equal to or greater than 60 min [ 36 ]. The following washing steps were defined using NaCl 5% HSA, allowing for higher SVF viability compared to RL, which was used in the Celution protocol, and ensuring an undetectable residual quantity of collagenase.…”

Inter-center comparison of good manufacturing practices-compliant stromal vascular fraction and proposal for release acceptance criteria: a review of 364 productions

“…Indeed, it is now well established that technical settings associated to AT processing can influence SVF characteristics and efficacy. They include both conditions of AT harvesting such as the harvesting area [ 5 ], the harvesting technique [ 6 , 7 ], and conditions of manufacturing [ 8 , 9 ] according to whether they are manual or automated, enzymatic, or mechanic [ 10 – 14 ]. Furthermore, flow cytometry is classically used to define the distribution of the main nucleated cell subsets within isolated SVF, as the best approach to approximate the SVF composition.…”

Inter-center comparison of good-manufacturing-practices compliant stromal vascular fraction and proposal for release acceptance criteria: a review of 364 productions

“…Indeed, it is now well established that technical settings associated to AT processing can in uence SVF characteristics and e cacy. They include both conditions of AT harvesting such as the harvesting area [5], the harvesting technique [6,7] and conditions of manufacturing [8,9] according to whether they are manual or automated, enzymatic or mechanic [10][11][12][13][14]. Furthermore, ow cytometry is classically used to de ne the distribution of the main nucleated cell subsets within isolated SVF, as the best approach to approximate the SVF composition.…”