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Toehold-mediated strand displacement reaction for dual-signal electrochemical assay of apolipoprotein E genotyping.

Apolipoprotein E (apoE) polymorphic genes are one of the main genetic determinants of Alzheimer's disease (AD) risk. Relying on the toehold-mediated strand displacement reaction, dual-signal electrochemical assay of apoE genotyping with potential applications in early diagnosis of AD has been achieved. The replacement of the surface-confined methylene blue (MB)- and ferrocene (Fc)-capped detection probe-modified AuNPs by the complementary sequences (Tc 1 and Tc 2, fragment of allele ε4 at codon 112 and that of allele ε3 or ε4 at codon 158, respectively), triggered by the highly specific strand displacement reaction, results in decreased voltammetric signals. In contrast, partial strand replacement caused by the single mismatched sequence (Tsm 1 and Tsm 2, fragment of allele ε2 or ε3 at codon 112 and that of allele ε2 at codon 158, respectively) produces larger voltammetric signals. The proposed method serves as a versatile platform for the discrimination of six apoE genotypes including three homozygotes (ε2/2, ε3/3, ε4/4) and three heterozygotes (ε2/3, ε2/4, ε3/4), and for the quantification of apoE ε3/3 from genomic DNA extracts of AD patients.