Preparation and Characterization of C-Reactive Protein Dual-Particle Latex-Enhanced Immunoturbidimetric Reagents

Objective and Impact Statement: This study aims to couple C-reactive protein (CRP) antibodies onto latex spheres of 2 different sizes to enhance the accuracy and sensitivity of CRP detection. Furthermore, it seeks to establish a robust methodological framework crucial for advancing the development of latex-enhanced immunoturbidimetric detection reagents. Introduction: CRP, an acute-phase protein, rapidly elevates in response to infections or tissue damage. Double-particle latex-enhanced immunoturbidimetry offers important advantages for accurately measuring CRP levels. Methods: CRP antibodies were coupled with 2 sizes of polystyrene latex spheres. Coupling rates were evaluated to determine optimal conditions. Particle sizes suitable for CRP detection, as well as coupling and mixing ratios, were optimized using automated biochemical analysis. Transmission electron microscopy and nanoparticle size analysis were employed to characterize the morphology and size changes of CRP antibodies and coupled latex spheres before and after immune reaction. Results: Optimization identified 168- and 80-nm latex sphere sizes, with CRP antibody coupling rates of 92% and 91%, respectively. The optimal ratios were 10:1.5 for large latex spheres to polyclonal antibodies and 5:1.5 for small latex spheres to monoclonal antibodies. A 1:8 mixing ratio of large to small latex spheres was effective. Transmission electron microscopy confirmed uniform sizes postcoupling, maintaining dispersion with no morphological changes. CRP reacted with the double-particle latex reagent, forming immune complexes that exhibited agglutination. Mixed latex spheres showed varied agglutination states with CRP concentration, altering solution absorbance. Conclusion: This study validates the efficacy of the dual-particle-size CRP antibody latex reagent, highlighting its potential for future immunoturbidimetric analysis applications.