Joris van Dongen

53 Comparison of SVF isolation procedures: a systematic review cells (CD31min) 7 . Three important subpopulations of the stromal cell population (CD45min/CD31min) are supra-adventitial cells: CD34pos/CD146min, pericytes: CD34pos/min/CD146pos and ASCs: CD34pos/CD90pos/CD105low 7,11,12,82 . Supra- adventitial cells and pericytes are both identified as precursor cells of ASCs, although there remains some controversyabout this item 11,12,80,83 . Ideally, to discriminate between those three cell types within the CD45min/CD31min subpopulation, CD146 and/or CD90 markers should be used additionally. However, in most studies two CD markers or inappropriate combinations of CD markers have been used to determine cell types; only Lin et al. used all the aforementioned combinations 73 . Because Lin et al. focus mainly on blood derived cells and not on the stromal cell population or pericytes, this did not affect their results. Doi et al. ascribed CD31min/CD34min/CD45min to the pericyte population, so therefore the CD34pos pericytes will be missed 76 . SundarRaj et al. and Güven et al. used CD34pos/CD31min to determine the number of ASCs 69,75 , while pericytes and supra-adventitial cells also express CD34. Therefore, the number of ASCs contains pericytes and supra-adventitial cells as well 7,11 . To cover pericytes, supra-adventitial cells and ASCs, Domenis et al., Aronowitz et al. and Mashiko et al. used CD34pos/CD31min/CD45min to determine the stromal cell population 66,70,72,74 . CD34pos is frequently used as a marker to describe cells with stem cell characteristics in both hematopoietic and non-hematopoietic stem cells 84 . The differences in use of CD marker expression to determine pericytes and the stromal cell population might be a possible explanation for the large variations found in SVF between different studies. No solid conclusions could be made about which isolation procedure generates the most stromal cells or pericytes. Unfortunately, a limited number of commercially available intraoperative SVF isolation procedures not yet have reached scientific validation at an acceptable level. The American Society for Aesthetic Plastic Surgery (ASAPS) and the American Society of Plastic Surgeons (ASPS) published a position statement in 2012 on fat grafting and stem cells 85 . All specialized equipment for the use of stem cell extraction should be fully verified regarding efficacy and safety before use in clinical settings. In 2013, the IFATS and ICTS proposed guidelines with standardized endpoints and methods to verify and compare SVF isolation procedures 5 . None of the included studies fully verified their isolation procedure according to these IFATS and ICTS guidelines. Moreover, viability was measured in different ways among studies ( e.g. directly on obtained SVF or after an extra non-intraoperative isolation protocol) and lipoaspirate was processed differently prior to isolation ( e.g. centrifugation or decantation). For those reasons, we propose new adjusted IFATS and ICTS guidelines to validate intraoperative isolation procedures (Fig. 3). All intraoperative isolation procedures should be validated using centrifuged adipose tissue to determine the actual volume of lipoaspirate prior to isolation. It is