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In vitro bioassay of endotoxin using fluorescein as a pH indicator in a macrophage cell culture system

 Dong Hee Lee  ;  Hak-Joon Sung  ;  Dong-Wook Han  ;  Min-Sub Lee  ;  Gyu Ha Ryu  ;  Maki Aihara  ;  Kosuke Takatori  ;  Jong-Chul Park 
 Yonsei Medical Journal, Vol.46(2) : 268-274, 2005 
Journal Title
 Yonsei Medical Journal 
Issue Date
Animals ; Biological Assay/methods* ; Biological Assay/standards ; Cells, Cultured ; Contrast Media* ; Culture Media/chemistry ; Endotoxins/analysis* ; Fluorescein* ; Hydrogen-Ion Concentration ; Limulus Test ; Lipopolysaccharides/analysis ; Macrophages/chemistry* ; Mice
Lipopolysaccharide ; macrophage ; fluorescein ; Limulus Amebocyte Lysate test ; Staphylococcus aureus ; pH
Based on the biological activity of endotoxin, we propose a possible new method for detecting endotoxin using a pH-indication system of macrophage culture media. After RAW 264.7 macrophage cells were treated with lipopolysaccharide (LPS), the addition of fluorescein to the LPS-treated media reproductively reduced its absorption and emission spectra (it was a dose-dependent reduction). The advantages of this LPS-detection method were compared with the Limulus Amebocyte Lysate (LAL) test by using purified bacterial LPS (Salmonella minnessota, Escherichia coli, and Pseudomonas aeruginosa). Additionally, the absorption and fluorescence intensity of fluorescein, following treatment of RAW 264.7 cells with a high concentration of Staphylococcus aureus (Gram-positive, lysed bacteria), could not generally be detected by the LAL test, but they were found to be reduced, in a dose-response relationship, with this new system. The macrophage culture system-method might be a good supplement to the LAL assay for detection of LPS, Gram-negative and Gram-positive bacteria.
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1. College of Medicine (의과대학) > Dept. of Medical Engineering (의학공학교실) > 1. Journal Papers
1. College of Medicine (의과대학) > Yonsei Biomedical Research Center (연세의생명연구원) > 1. Journal Papers
Yonsei Authors
Park, Jong Chul(박종철) ORCID logo https://orcid.org/0000-0003-0083-5991
Sung, Hak-Joon(성학준) ORCID logo https://orcid.org/0000-0003-2312-2484
Lee, Dong Hee(이동희)
Lee, Min Sub(이민섭)
Han, Dong Wook(한동욱)
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