Open AccessFlow cytometric analysis of Xenopus laevis and X. tropicalis blood cells using acridine orange
Author(s) -
Koji Sato,
Azusa Uehara,
Sayaka Kinoshita,
Ikki Nomura,
Minami Yagi,
Yuta Tanizaki,
Yu Matsuda-shoji,
Atsushi Matsubayashi,
Nobuyasu Endo,
Yutaka Nagai,
Takashi Kato
Publication year - 2018
Publication title -
scientific reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.24
H-Index - 213
ISSN - 2045-2322
DOI - 10.1038/s41598-018-34631-0
Subject(s) - acridine orange , xenopus , biology , flow cytometry , staining , microbiology and biotechnology , red blood cell , blood cell , cell , fluorescence , orange (colour) , genetics , biochemistry , gene , physics , quantum mechanics , horticulture
Automated blood cell counters can distinguish cells based on their size and the presence or absence of a nucleus. However, most vertebrates have nucleated blood cells that cannot be counted automatically. We established an alternative automatic method for counting peripheral blood cells by staining cells with the fluorescent dye acridine orange (AO) and analysing cell populations using flow cytometry (FCM). As promising new animal models, we chose Xenopus laevis and three inbred strains of X. tropicalis . We compared the haematological phenotypes, including blood cell types, cell sizes, cellular structure, and erythrocyte lifespans/turnover rate among X. laevis and the three inbred strains of X. tropicalis . Each cell type from X. laevis was sorted according to six parameters: forward- and side-scattered light emission, AO red and green fluorescence intensity, and cellular red and green fluorescence. Remarkably, the erythrocyte count was the highest in the Golden line, suggesting that genetic factors were associated with the blood cells. Furthermore, immature erythrocytes in anaemic X. laevis could be separated from normal blood cells based on red fluorescence intensity. These results show that FCM with AO staining allows for an accurate analysis of peripheral blood cells from various species.