Accuracy and Reproducibility of Stem Cell Side Population Measurements on Clinically Relevant Products

ISSN: 2212-3946 (Online)
ISSN: 1574-888X (Print)


Volume 9, 6 Issues, 2014


Download PDF Flyer




Current Stem Cell Research & Therapy

Aims & ScopeAbstracted/Indexed in


Submit Abstracts Online Submit Manuscripts Online

Editor-in-Chief:
Anthony Atala
Wake Forest University School of Medicine,
Medical Center Boulevard
Winston Salem, NC 27157
USA


View Full Editorial Board

Subscribe Purchase Articles Order Reprints

Current: 2.861
5 - Year: 3.00

Accuracy and Reproducibility of Stem Cell Side Population Measurements on Clinically Relevant Products

Author(s): Ariadna Avendano, Irene Sales-Pardo, Maria Dolores García-Godoy, Laura G. Rico, Pedro Marin and Jordi Petriz

Affiliation: Laboratori 207, Vall d'Hebron Institut de Recerca, Pº Vall d'Hebron 119-129, 08035 Barcelona; Spain.

Abstract

Introduction: In 1996, Goodell et al. first described a rare subpopulation of bone marrow stem cells termed the Side Population (SP). SP cells are known to be CD34 negative and to have a high repopulating capability. The SP was identified by ultraviolet excitation based on the efflux of the DNA binding dye, Hoechst 33342 (Ho342). ABCG2, a halftransporter that belongs to the ATP binding cassette transporter superfamily, is the major contributor to the SP phenotype by actively pumping Ho432 selectively from stem cells. To date, very little is known about the identification of the SP in peripheral blood samples, and about its peripheral circulation, enrichment or isolation to evaluate its therapeutic potential. Due to the SP potential role in tissue regeneration, we studied the numbers of the SP in bone marrow and peripheral blood samples in regard to count accuracy and reproducibility. Materials and methods: Bone marrow (BM) and apheresis (AP) specimens were obtained from healthy donors and patients undergoing stem cell transplantation. Bone marrow samples were obtained by aspiration. Peripheral blood cells after granulocyte colony stimulating factor (G-CSF) mobilization with or without chemotherapy, were obtained by apheresis. All samples were prepared for identification of SP cells by flow cytometry. Results: SP cells were detected in only 19 of 111 apheretic products, with relative frequency ranging from 0.01 to 4.75% of cells by the Ho342 exclusion method and flow cytometry analysis. Cell preparations used for these measurements consisted of 5 x 106 cells. However, no SP cells were detected when aliquots from the same positive specimens, consisting of previously stained 55 x 106 cells and fractionated into independent aliquots with 5 x 106 cells were used. Conclusions: In this study, we show that there is great variability in SP cell numbers when aliquots obtained either from leukapheresis or bone marrow products represent about 1% of the total product volume. In contrast, when aliquots represented about 12% of the total product volume SP cells measurements were consistent. The high cell number of some specimens can be a limitation for the accurate identification and isolation of the SP compartment. Aliquots containing a minimum of 55 × 106 cells should be used for statistical significance.

Keywords: ABCG2, CD34, flow cytometry, side population, stem cell.

Purchase Online Rights and Permissions

  
  



Article Details

Volume: 9
Issue Number: 6
First Page: 526
Last Page: 534
Page Count: 9
DOI: 10.2174/1574888X09666140425110424
Advertisement

Related Journals




Webmaster Contact: urooj@benthamscience.org Copyright © 2014 Bentham Science