Development and validation of a novel latex particle-enhanced turbidimetric immunoassay for fecal calprotectin.

BACKGROUND

Fecal calprotectin (FCP) is a crucial non-invasive biomarker for diagnosing and monitoring inflammatory bowel disease (IBD). However, the standard enzyme-linked immunosorbent assay (ELISA) is time-consuming and labor-intensive. This study aimed to develop a novel, independent latex particle-enhanced turbidimetric immunoassay (PETIA) suitable for open-channel automated biochemical analyzers, offering a flexible and cost-effective alternative to existing commercial PETIA kits.

METHODS

Recombinant human S100A8 and S100A9 proteins (the subunits of calprotectin) were expressed in E. coli and purified. Polyclonal antibodies were generated in rabbits by immunization with these proteins. The antibodies were then covalently coupled to carboxylated latex particles (188 nm) to create the PETIA reagent. The assay was validated on an LC-400 specific protein analyzer. Analytical performance-including repeatability, within-lab precision, and interference-was assessed. The method was evaluated using clinical fecal samples (n = 104) and compared with a commercial ELISA kit (BÜHLMANN fCAL).

RESULTS

The developed PETIA demonstrated a measuring range of 0-1500 µg/g based on a six-point calibration curve (R2 = 0.99). A strong correlation was observed between the new PETIA and the reference ELISA (r = 0.98). The regression equation was y = 1.04x + 8.82. Bland-Altman analysis confirmed good agreement between the two methods, with 95.2% of data points within the 95% limits of agreement.

CONCLUSION

The PETIA showed strong correlation with the established ELISA for FCP quantification, offering a rapid, automated alternative suitable for high-throughput laboratories. This study, encompassing development from antibody to reagent, supports its translational potential.