Open sandwich fluorescence-linked immunosorbent assay for detection of soy isoflavone glycosides

• Main Authors: Seiichi Sakamoto, Hiroki Uchiyama, Gorawit Yusakul, Napat Kyokong, Benyakan Pongkitwitoon, Waraporn Putalun, Hiroyuki Tanaka, Satoshi Morimoto
• Other Authors: Chulalongkorn University
• Format: Article
• Published: 2022
• Subjects: Agricultural and Biological Sciences; Chemistry
• Online Access: https://repository.li.mahidol.ac.th/handle/123456789/75561
• Institution: Mahidol University

To detect major soy isoflavone glycosides, namely daidzin (DZ) and genistin (GEN), novel open sandwich fluorescence-linked immunosorbent assay (os-FLISA) was developed by taking advantage of enhanced interactions between variable regions of heavy (VH) and light chain (VL) domains in the presence of an antigen. The VH and VL genes were expressed in Escherichia coli as a chimera protein with green fluorescence protein (AcGFP1) and maltose-binding protein (MBP), respectively. Comprehensive characterization of os-FLISA displayed nearly the same specificity as parental DZ- and GEN-specific monoclonal antibody, demonstrating the potential of the developed assay for detection of both DZ and GEN. Their detectable range in this system exhibited at 0.1–12.5 μg mL−1. Subsequent validation analysis revealed that os-FLISA was reliable and accurate system for detection of total soy isoflavone glycosides. Notably, this is the first FLISA based on an open sandwich system, which can be employed for the detection of small molecules.