Label-Free and Separation-Free Atomic Fluorescence Spectrometry-Based Bioassay: Sensitive Determination of Single-Strand DNA, Protein, and Double-Strand DNA.

Based on selective and sensitive determination of Hg(2+) released from mercury complex by cold vapor generation (CVG) atomic fluorescence spectrometry (AFS) using SnCl2 as a reductant, a novel label-free and separation-free strategy was proposed for DNA and protein bioassay. To construct the DNA bioassay platform, an Hg(2+)-mediated molecular beacon (hairpin) without labeling but possessing several thymine (T) bases at both ends was employed as the probe. It is well-known that Hg(2+) could trigger the formation of the hairpin structure through T-Hg(2+)-T connection. In the presence of a specific target, the hairpin structure could be broken and the captured Hg(2+) was released. Interestingly, it was found that SnCl2 could selectively reduce only free Hg(2+) to Hg(0) vapor in the presence of T-Hg(2+)-T complex, which could be separated from sample matrices for sensitive AFS detection. Three different types of analyte, namely, single-strand DNA (ssDNA), protein, and double-strand DNA (dsDNA), were investigated as the target analytes. Under the optimized conditions, this bioassay provided high sensitivity for ssDNA, protein, and dsDNA determination with the limits of detection as low as 0.2, 0.08, and 0.3 nM and the linear dynamic ranges of 10-150, 5-175, and 1-250 nM, respectively. The analytical performance for these analytes compares favorably with those by previously reported methods, demonstrating the potential usefulness and versatility of this new AFS-based bioassay. Moreover, the bioassay retains advantages of simplicity, cost-effectiveness, and sensitivity compared to most of the conventional methods.