title: Evaluation of 2 Purification Methods for Isolation of Human Adipose-Derived Stem Cells Based on Red Blood Cell Lysis With Ammonium Chloride and Hypotonic Sodium Chloride Solution.
abstract: BACKGROUND
  The present study was conducted to compare 2 purification methods for isolation of human adipose-derived stromal vascular fraction or stem cells (ADSCs) based on red blood cell (RBC) lysis with 155 mM ammonium chloride (NH4Cl) and hypotonic sodium chloride (NaCl) solution, and try to develop a safe, convenient, and cost-effective purification method for clinical applications.
  
  
  METHODS
  Adipose-derived stem cells and RBC were harvested from the fatty and fluid portions of liposuction aspirates, respectively. The suitable concentration of hypotonic NaCl solution on RBC lysis for purification of ADSCs was developed by RBC osmotic fragility test and flow cytometry analysis. The effects of 155 mM NH4Cl or 0.3% NaCl solution on ADSCs proliferation and RBC lysis efficiency were examined by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide assay and lysis efficiency test, respectively. In addition, the adipogenic and osteogenic capabilities, phenotype and genetic stability of ADSCs were evaluated by oil red staining, alkaline phosphatase activity measurement, flow cytometry, and karyotype analysis, respectively.
  
  
  RESULTS
  Sodium chloride solution in 0.3% concentration effectively removed RBCs and did not influence the survival of ADSCs in the 10-minute incubation time. The lysis efficiency did not differ significantly between 0.3% NaCl and 155 mM NH4Cl. Moreover, the adipogenic and osteogenic capabilities, surface marker expression and karyotype of the ADSCs were not affected by lysis solutions or by lysis per se. However, the proliferation capacity in the 0.3% NaCl group was superior to that in 155 mM NH4Cl group.
  
  
  CONCLUSIONS
  Our data suggest that 0.3% NaCl solution is useful for isolating ADSCs from liposuction aspirate for clinical applications with safety, convenience, and cost-effect.
authors.affiliation: From the *Department of Plastic Surgery, The First Affiliated Hospital of Jinan University, Key Laboratory of Regenerative Medicine, Ministry of Education, Guangzhou; †Department of Plastic Surgery, Guangzhou General Hospital of Guangzhou Military Command, Guangdong Province; and ‡Department of Plastic Surgery, Changhai Hospital, The Second Military Medical University, Shanghai, P.R. China.
authors.forename: Sheng-Hong
authors.initials: SH
authors.lastname: Li
authors.forename: Xuan
authors.initials: X
authors.lastname: Liao
authors.forename: Tian-En
authors.initials: TE
authors.lastname: Zhou
authors.forename: Li-Ling
authors.initials: LL
authors.lastname: Xiao
authors.forename: Yuan-Wen
authors.initials: YW
authors.lastname: Chen
authors.forename: Fan
authors.initials: F
authors.lastname: Wu
authors.forename: Jing-Ru
authors.initials: JR
authors.lastname: Wang
authors.forename: Biao
authors.initials: B
authors.lastname: Cheng
authors.forename: Jian-Xing
authors.initials: JX
authors.lastname: Song
authors.forename: Hong-Wei
authors.initials: HW
authors.lastname: Liu
mesh_terms: D000328:Adult; D000643:Ammonium Chloride; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods*; D005260:Female; D006461:Hemolysis; D006801:Humans; D007038:Hypotonic Solutions; D015187:Lipectomy; D008297:Male; D059630:Mesenchymal Stem Cells / Q000502:physiology; D008875:Middle Aged; D012965:Sodium Chloride; D050507:Subcutaneous Fat, Abdominal / Q000166:cytology* / Q000601:surgery
chemical_list: D007038:Hypotonic Solutions; D000643:Ammonium Chloride; D012965:Sodium Chloride