[A fluorimetric microplate assay for detecting diarrheic shellfish poisoning toxins].
OBJECTIVE
To establish a new assay for detecting the cytotoxicity of diarrheic shellfish poisoning (DSP) toxins. The assay is based on the depolymerization of F-actin induced by okadaic acid (OA), which is one of main components of DSP toxins.


METHODS
HL-7702 Liver cells were stained with Oregon Green-514 phalloidin as a fluorescent marker for F-actin. The change of the fluorescence of F-actin treated with OA was measured with a fluorimetric microplate reader. The detection results of this assay and ELISA were compared to evaluate the reliability of the assay.


RESULTS
OA caused an increase of F-actin depolymerization in a dose-dependent manner. There was a linear relationship between the concentration of OA and the depolymerization of F-actin in the range of 2.5 - 40 nmol/L of OA in 24 hours (R2 = 0.993). The detection limit of the F-actin fluorescence assay for OA was 2.01 microg/100g muscles in shellfish extracts, and the recoveries were 90% - 100%. The results of the fluorescence assay were consistent with other methods mentioned above (R2 = 0.830).


CONCLUSIONS
F-actin fluorescence assay was a promising method for the detection of OA in shellfish for its convenience, shortcut and sensitivity.
Toxicology Laboratory, Shenzhen Center for Disease Control and Prevention, Shenzhen 518055, China. hhy424@126.com
Haiyan
H
Huang
Aijun
A
Huang
Yingbin
Y
Fu
Wei
W
Huang
Xintian
X
Yu
Jianjun
J
Liu
D000199:Actins; D001681:Biological Assay; D003967:Diarrhea / Q000209:etiology; D004797:Enzyme-Linked Immunosorbent Assay; D005452:Fluoresceins; D005470:Fluorometry; D006801:Humans; D057230:Limit of Detection; D019319:Okadaic Acid; D015203:Reproducibility of Results; D012758:Shellfish; D057096:Shellfish Poisoning; D014118:Toxins, Biological / Q000032:analysis*
D000199:Actins; D005452:Fluoresceins; D014118:Toxins, Biological; C401939:oregon green 514; D019319:Okadaic Acid