Rapid bead-based immunoassay for measurement of mannose-binding lectin.

ABSTRACT

Mannose-binding lectin (MBL) is a serum protein, which functions as an opsonin and initiator of the lectin pathway of complement. The serum concentration of MBL shows great interindividual variation because of common polymorphisms in the MBL2 gene. Although several quantitative MBL immunoassays have been developed more automated platforms for MBL analysis is urgently needed. To pursue this, we set out to develop a flexible bead-based MBL immunoassay. Serum was obtained from 98 healthy individuals and 50 patients investigated for possible immunodeficiencies. We used the Luminex xMAP bead array technology employing a mouse monoclonal anti-MBL antibody both for coating and detection. The assay was fast and reliable for measurements of MBL concentrations both in the lower and upper range. The lower detection limit was found to be 6.5 microg/l. The intra-assay coefficient and the interassay coefficient were found be 7.88% and 5.70%, respectively. A close correlation between the new assay and a reference MBL measurement ELISA was found (rho 0.9381, P < 0.0001). The bead-based assay was less sensitive to interfering anti-murine antibodies in the blood samples than when the antibodies employed were used in the reference polystyrene-based ELISA. The new assay could be performed in 3 h with less than 25 microl serum required of each sample. These results show that MBL can be measured readily using a bead-based platform, which may form an efficient basis for a multiplex approach to measure different antigens in the same sample.