New latex agglutination assay for the determination of lactoferrin in human milk

Lactoferrin (LF) in human milk has various biological properties and contributes to the prevention of preterm birth complications. Enzyme-linked immunosorbent assay (ELISA) is one of the most commonly used methods to measure LF in human milk, but this method is time-consuming and laborious. In Japanese human milk banks, the concentration of LF in donor human milk (DHM) is measured routinely. Here, we reported a rapid, simple, and accurate method for determining LF in human milk using a new reagent based on a latex agglutination assay. We obtained 208 human milk pools from 148 mothers, and samples were collected before and after Holder pasteurization. Milk samples were diluted 100- or 200-fold and LF concentrations were measured by a latex agglutination assay using an automated analyzer. The reagent was validated in terms of repeatability, linearity, detection limit, recovery, and comparison with ELISA. The coefficient of variation (CV) for intra-assay precision ranged from 0.6 to 5.0% in human milk with high, medium, and low LF concentrations. The linearity was also tested by serial sample dilution and was confirmed up to 16 µg/mL with a detection limit of 0.2 µg/mL. The recovery rates in a spiked recovery test were ranged from 90 to 120% at high, medium, and low concentrations of LF. Furthermore, a strong correlation was observed between LF levels determined by the latex agglutination assay and ELISA (r = 0.978, p