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Novel DRAQ5™/SYTOX® Blue Based Flow Cytometric Strategy to Identify and Characterize Stem Cells in Human Breast Milk
Author(s) -
Keller Titus,
Wengenroth Leonie,
Smorra Denise,
Probst Kristina,
Kurian Leo,
Kribs Angela,
Brachvogel Bent
Publication year - 2019
Publication title -
cytometry part b: clinical cytometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.646
H-Index - 61
eISSN - 1552-4957
pISSN - 1552-4949
DOI - 10.1002/cyto.b.21748
Subject(s) - flow cytometry , breast milk , stem cell , biology , cytometry , andrology , immunology , microbiology and biotechnology , medicine , biochemistry
Background Human breast milk could be an important stem cell source for the development of newborn and preterm infants, but quantitative data on the stem cell content in breast milk at various gestational stages are needed to determine the clinical value of breast milk as a source of stem cells. Breast milk also contains milk fat globules, lipid droplets of different sizes, debris and dead cells and these components hamper flow cytometry analysis of human breast milk samples. Methods Here, we originally used standard protocols for flow cytometry to characterize cell populations in human breast milk but failed to discriminate between cells and noncellular components. We then applied a centrifugation protocol to separate cream and skim milk from the cell‐containing pellet and used a novel staining protocol with DRAQ5™ and SYTOX® blue dye as well as antibodies to characterize cells within the pellet fraction. Results Flow cytometry analysis identified viable DRAQ5™ + /SYTOX® Blue − cells and determined the content of CD11b + monocytes and TRA‐1‐81 + putative stem cells in human breast milk samples. Conclusions Hence, we developed a novel and reliable flow cytometry based‐approach to quantify subpopulation of cells in human breast milk with a high content of milk fat globules, lipid droplets, and particles. This approach will improve the identification and quantification of breast milk cells and allow standardizing the flow cytometry‐based evaluation of the stem cell content. © 2018 International Clinical Cytometry Society