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An automated, single microbead-arrayed μ-fluidic immunoassay (AMIA) device is innovatively devised in this study, which enables the highly sensitive and simultaneous detection of multiplex biomarkers with fully automatic operations. The AMIA platform not only achieves automated assay processing and multiplexed target detection by integrating single microbead manipulation, sample loading, multistep washing, and immunoreaction on a microfluidic chip but also confers high sensitivity due to the highly efficient signal enriching effect on a single microbead by the use of only a routine sandwich immunoreaction. As such, as low as the pg/mL level of multiplexed protein biomarkers can be simultaneously determined in a quite small volume of serum (∼20 μL is enough), which can well meet the clinical demand for disease screening and prognosis. What is more, the detection results of several clinically important biomarkers in clinical samples with the AMIA platform exhibit excellent consistency with those obtained by using a standard clinical test. Thus, in virtue of the excellent features in terms of high sensitivity, multiplexing capability, generality, and high degree of automation, the AMIA provides a practical and user-friendly platform for assaying different biomarkers in clinical diagnostics and point-of-care testing.

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