Conventional immunochemical methods used in clinical analysis are often not sensitive enough for early-stage diagnosis, resulting in the need for novel assay formats. Here, we provide a detailed comparison of the effect of different labels and solid supports on the performance of heterogeneous immunoassays. When comparing three types of streptavidin-modified labels─horseradish peroxidase, carboxyfluorescein, and photon-upconversion nanoparticles (UCNPs)─UCNPs led to the most sensitive and robust detection of the cancer biomarker prostate-specific antigen. Additionally, we compared the immunoassay formats based on conventional microtiter plates and magnetic microbeads (MBs). In both cases, the highest signal-to-background ratios and the lowest limits of detection (LODs) were obtained by using the UCNP labels. The MB-based upconversion-linked immunosorbent assay carried out with a preconcentration step provided the lowest LOD of 0.46 pg/mL in serum. The results demonstrate that the use of UCNPs and MBs can significantly improve the sensitivity and working range of heterogeneous immunoassays for biomarker detection.

CEITEC MU Masaryk University Kamenice 5 625 00 Brno Czech Republic

Department of Biochemistry Faculty of Science Masaryk University Kamenice 5 625 00 Brno Czech Republic

Institute of Analytical Chemistry Chemo and Biosensors University of Regensburg 93053 Regensburg Germany

Institute of Analytical Chemistry of the Czech Academy of Sciences Veveří 967 602 00 Brno Czech Republic

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