Development of a quantitative detection card for heart-type fatty acid-binding protein based on background fluorescence quenching immune chromatography

Abstract

Background: To establish a fast and simple quantitative method for detection of heart-type fatty acid-binding protein (H-FABP) in serum based on a background fluorescence quenching im munochromatographic assay. Methods: A detection card based on the double-antibody sandwich double-antibody method with background fluorescence quenching was developed for quantitative m easurement of H-FABP in serum. The optimal concentrations of control for coating the test and control lines were determined as well as the concentrations of gold-labeled antibodies used in preparing the detection system. The detection method for H-FABP in serum was established and validated using real-world clinical samples. Results: The optimal concentrations of labeling antibody and coating antibody were 5.0 mg/mL and 1.0 mg/m L, respectively. The test card had a sensitivity of 1.15 ng/m L over a linear concentration range of 0-100 ng/m L. Based on three batches prepared for testing the card, the relative standard deviation (RSD) within batches was less than 15% without a significant difference (P = 0.942 ). The detection method was tested against common interfering substances in serum, such as bilirubin, triglyceride and serum anticoagulants ethylenediamine tetraacetic acid (EDTA), heparin, and sodium citrate, and no significant cross-reaction was detected. The test method was further validated with 50 clinical serum samples, and the test results were COM parable with standard reference detection methods with good correlation (R = 0.95 ). Conclusion: O ur study presents a new method with strong specificity and sensitivity for the detection of H-FABP in serum, which could promote H-FABP detection in a broad range of applications.

Keywords

• heart-type fatty acid binding protein
• background fluorescence quenching
• immunochromatography