Immunomagnetic separation using carbonyl iron powder and flow cytometry for rapid detection of <Emphasis Type="Italic">Flavobacterium psychrophilum</Emphasis> |
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Authors: | Kyoko Hibi Hideki Ushio Hideo Fukuda Kohji Mitsubayashi Tetsuhito Hayashi Huifeng Ren Hideaki Endo |
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Institution: | (1) Faculty of Marine Science, Tokyo University of Marine Science and Technology, 4–5–7, Konan, Minato-ku Tokyo, 108–8477, Japan;(2) Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, 2–3–10, Kanda-Surugadai, Chiyoda-ku Tokyo, 101–0062, Japan |
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Abstract: | Bacterial cold water disease, caused by Flavobacterium psychrophilum, is a serious problem in the aquaculture industry worldwide. Several methods to prevent and treat cold water disease have
been studied. Although detection at the early stage of F. psychrophilum infection is very important for the prevention and treatment of cold water disease, an effective detection method has not
yet been developed. The use of flow cytometry (FCM) for the rapid determination of bacterial cell numbers with high sensitivity
is beginning to attract attention. Immunomagnetic separation (IMS) has also been used to detect F. psychrophilum. The purpose of the present study was to develop a method to quickly determine the number of bacterial cells by combining
the FCM and IMS methods. Because samples can be more effectively concentrated using smaller magnetic beads and stronger magnetism,
we used carbonyl iron powder as the magnetic beads for the IMS. The detection level of F. psychrophilum using FCM combined with IMS was 5 orders lower than that using FCM without IMS. The values determined using FCM combined
with IMS strongly correlated with those obtained using the colony-counting method, in the range of approximately 10–108 colony-forming units per milliliter. One FCM assay could be completed within 60 s and the total assay time, including sample
preparation, was less than 2 h. The combined method of FCM with IMS developed in this study can be used reliably for the rapid
detection of F. psychrophilum. |
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Keywords: | Bacteria Immunomagnetic separation Magnet beads Carbonyl iron powder Flow cytometry Bacterial cold water disease |
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