High-throughput assay for detection of soybean lipoxygenase-1.
A high-throughput assay was developed to detect soybean lipoxygenase 1 (LOX-1) using a multilabel plate reader. The assay was also adapted to a single cell fluorometer. Fluorescein is degraded by linoleic hydroperoxide produced from soybean lipoxygenase and linoleic acid. The decrease in fluorescence is measured over time, and the area-under-the-curve (AUC) is used to quantify the LOX-1 content of soybean extract. A dose-dependent response is seen with varied dilutions of pure LOX enzyme or soybean extracts. Percent recovery was between 97% and 108%, and relative standard deviation was 4.3%. Advantages of the assay include the reduced preparation time of samples and reduced use of reagents in the high-throughput assay. Multiple samples can be measured in a single run with a multilabel plate reader.
Department of Nutrition and Food Science, University of Maryland, College Park, Maryland 20742, United States.
Monica
M
Whent
Tony
T
Ping
William
W
Kenworthy
Liangli Lucy
LL
Yu
D057075:Enzyme Assays / Q000379:methods*; D057166:High-Throughput Screening Assays / Q000379:methods*; D008084:Lipoxygenase / Q000032:analysis* / Q000378:metabolism; D010940:Plant Proteins / Q000032:analysis* / Q000378:metabolism; D013025:Glycine max / Q000737:chemistry / Q000201:enzymology*
D010940:Plant Proteins; C101084:lipoxygenase L-1; D008084:Lipoxygenase