A Semi-quantitative Lateral Flow Immunoassay for Detection of an Osteoarthritis Biomarker in Human Synovial Fluid
Jang, Yo Han; Park, Yoo Min; Kim, Su Jin; Yoon, Hyun C.
Institute for Medical Sciences, Ajou University School of Medicine
We developed a reading device-less semi-quantitative lateral immunoassay (SQ-LFI) method to measure the concentration of cartilage oligomeric matrix protein (COMP) in human synovial fluid (SF) for the osteoarthritis (OA) diagnosis. The COMP is one of the important cartilage degradation biomarker of OA diagnosis. Despite the good selectivity and operation convenience of conventional lateral flow immunoassay (LFI), it provides only a qualification signal (ON/OFF). Although there are some quantifiable LFI systems that provide quantification signal, these systems require external signal reading device for the quantification. To overcome these drawbacks and develop semi quantifiable LFI systems which require no external device, immobilization pattern of biomarker-capturing antibodies were converted from single line-type into herringbone-type arrayed spot. The COMP capture antibody (16F12) was spotted in type of herringbone-pattern on the test zone of nitrocellulose membrane (25 spots) and the COMP detection antibody (12C4) was conjugated with the gold-nanoparticles (AuNPs) to obtain the optical signal. The capture antibody and detection antibody are specifically recognizing and binding to different site of COMP molecules. The COMP in human synovial fluid binds to AuNP-Ab and then COMP/AuNP-Ab complex were captured by immobilized capture antibody spots on the membrane. Contrast to the test result of conventional LFI visualized as single colored-line, the test result of SQ-LFI appeared as bunch of colored-spots. The number of colored spot in the test zone was increased in proportion to the concentration of biomarker. By the simple counting of developed spots on the test zone, a semi-quantitative analysis of COMP was easily accomplished without any external device. A reliable calibration curve from the developed immunoassay exhibited a range from 0.63 to 20 μg/mL of COMP in SF. The test results from SQ-LFI were compared with the results from conventional ELISA test and it showed a good correlation.
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