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for direct comparison of reported biological properties and experimental outcomes,
especially in the context of cell-based therapy (Dominici et al. 2006 )?
Dominici and colleagues suggested in 2006 that the standard isolation protocol
developed by Zuk and co-workers ( 2001 , 2002 ) should be accepted as an estab-
lished methodology to obtain SVF from raw lipoaspirate (Dominici et al. 2006 ).
Most research groups however make adaptations to this methodology, and this com-
plicates the comparison of results between groups. Previous studies have suggested
that ASCs exhibit an average population doubling time of 60 h or generally 2–4
days, depending on the donor’s age, the type (white or brown) and location (subcu-
taneous or visceral) of the adipose tissue, the type of surgical procedure, culture
conditions, growth factors, plating or seeding densities, passage number, and media
formulations (Fossett et al. 2012 ; Gimble et al. 2007 ; Mizuno 2009 ). This again
highlights the many factors to consider when developing standardized isolation
protocols.
Different fat processing techniques have also been evaluated. A prospective
cross-sectional study evaluated three widely used fat processing techniques in plas-
tic surgery for the viability and number of adipocytes and ASCs isolated from col-
lected lipoaspirate (Condé-Green et al. 2010 ). All samples were collected using the
established Coleman technique under regional anesthesia. The aspirate was pro-
cessed using three different techniques, namely, (1) decantation, (2) washing, and
(3) centrifugation. The three basic layers, the superior oily liquid supernatant, the
fi rmer white-yellow tissue, and the inferior layer consisting mostly of blood con-
taminants including the infi ltration and washing liquids, were identifi ed with all
three techniques. A fourth layer, the pellet, was identifi ed with centrifugation only.
Signifi cant differences were observed with regard to viable adipocytes in the middle
fi rm tissue layer between various processing techniques ( p = 0.0075), where cen-
trifugation rendered adipocytes nonviable compared to decantation and washing
techniques.
Flow cytometric analysis has revealed various differences in ASCs, hematopoi-
etic cells (blood contaminants), and endothelial cells, comparing the middle fi rm
tissue layers of all three different processing techniques and the pellet of the centri-
fuged samples (Condé-Green et al. 2010 ). The fi rm tissue layer of the decantation
process contained large amounts of blood contaminants and very few ASCs and
endothelial cells. The fi rm tissue layer of the washing process contained few blood
contaminants and more endothelial cells and ASCs, compared to the decantation
process. The fi rm tissue layer of the centrifuged samples contained the least number
of ASCs, blood contaminants, and endothelial cells, whereas the pellet of the cen-
trifuged samples contained the greatest number of ASCs, blood contaminants, and
endothelial cells. In addition, the fi rm tissue layer from the centrifuged samples did
not expand and proliferate in vitro, while the pellet of the centrifuged samples dem-
onstrated extensive proliferation and expansion (Condé-Green et al. 2010 ). A recent
study confi rmed this fi nding by comparing centrifuged and non-centrifuged sam-
ples collected from subcutaneous adipose tissue in the abdominal area using the
Coleman technique and revealed that the centrifuged samples contained a signifi -
cantly greater SVF and ASC yield (Iyyanki et al. 2015 ).
F.A. van Vollenstee et al.