Disease diagnosis typically requires to determine concentration of multiple biomarkers in patient serums. Here, a novel method for multiplex immunoassays is proposed and the feasibility is demonstrated. The method utilizes the differential affinity between aptamers and multiple analytes for multiplex immunoassays. During the selection, aptamers capable of binding to multiple analytes with different affinities are screened from a random oligonucleotide library using the MARAS procedure with different magnetic field conditions for different target analytes. During the detection, the same magnetic field conditions are applied to differentiate different target analytes in blind serums. The results show that the recovery rates of the spiked targets in BD buffer and blind serums are similar. Moreover, there is a minimal interference resulting from non-specific binding of molecules in serums other than the target molecules. Therefore, the use of differential affinities between aptamers and different analytes for multiplex immunoassays is proved to be feasible.
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