1. Combinatorial semisynthesis of biomarker-IgM complexes.
- Author
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Veggiani G, Zuin J, Beneduce L, Gallotta A, Pengo P, and Fassina G
- Subjects
- Animals, Antigen-Antibody Complex blood, Antigens, Neoplasm blood, Avidin, Biomarkers, Biomarkers, Tumor, Biotin, Carcinoma, Squamous Cell blood, Carcinoma, Squamous Cell immunology, Cattle, Enzyme-Linked Immunosorbent Assay, Humans, Immunoglobulin M blood, Immunoglobulin M isolation & purification, Antigen-Antibody Complex immunology, Antigens, Neoplasm immunology, Immunoglobulin M immunology
- Abstract
Circulating immune complexes formed by tumor antigens and immunoglobulin M (IgM) represent a novel class of biomarkers with diagnostic value for early cancer detection. The quantitative analysis of these immune complexes is achieved by enzyme-linked immunosorbent assay (ELISA) methods using a purified calibrator from samples of patients with cancer. These complexes obtained from samples of human origin are not suitable for cost-effective production processes with high safety standards. Given the ill-defined biomarker/IgM ratio in these complexes, semisynthesis with retention of functional properties is difficult to achieve and may vary widely according to the batch-to-batch heterogeneity of starting biological preparations. Here the authors describe the development of a combinatorial method for defining the optimal reaction conditions for the reproducible semisynthesis of biomarker-IgM complexes by exploiting the biotin-avidin technology. The method relies on screening by ELISA the 3D composition space defined by the combinatorial variation of biotinylated-biomarker, biotinylated-IgM, and avidin concentrations aiming to select those conditions leading to biomarker-IgM complexes with the highest immunoreactivity. The method allows the reproducible synthesis of species with immunoreactivity comparable to that of natural immune complexes and endowed with sufficient stability to be used as calibrators in ELISA.
- Published
- 2010
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