Label-free detection of biomolecules using inductively coupled plasma mass spectrometry (ICP-MS).

Bioassays using inductively coupled plasma mass spectrometry (ICP-MS) have gained increasing attention because of the high sensitivity of ICP-MS and the various strategies of labeling biomolecules with detectable metal tags. The classic strategy to tag the target biomolecules is through direct antibody-antigen interaction and DNA hybridization, and requires the separation of the bound from the unbound tags. Label-free ICP-MS techniques for biomolecular assays do not require direct labeling: they generate detectable metal ions indirectly from specific biomolecular reactions, such as enzymatic cleavage. Here, we highlight the development of three main strategies of label-free ICP-MS assays for biomolecules: (1) enzymatic cleavage of metal-labeled substrates, (2) release of immobilized metal ions from the DNA backbone, and (3) nucleic acid amplification-assisted aggregation and release of metal tags to achieve amplified detection. We briefly describe the fundamental basis of these label-free ICP-MS assays and discuss the benefits and drawbacks of various designs. Future research is needed to reduce non-specific adsorption and minimize background and interference. Analytical innovations are also required to confront challenges faced by in vivo applications.

Arsenic species in electronic cigarettes: Determination and potential health risk.

Human exposure to contaminants from electronic cigarettes (e-cigarettes) and the associated health effects are poorly understood. There has been no report on the speciation of arsenic in e-liquid (solution used for e-cigarettes) and aerosols. We report here determination of arsenic species in e-liquids and aerosols generated from vaping the e-liquid. Seventeen e-liquid samples of major brands, purchased from local and online stores in Canada and China, were analyzed for arsenic species using high-performance liquid chromatography and inductively coupled plasma mass spectrometry. Aerosols condensed from vaping the e-liquids were also analyzed and compared for arsenic species. Six arsenic species were detected, including inorganic arsenate (iAsV), arsenite (iAsIII), monomethylarsonic acid (MMA), and three new arsenic species not reported previously. In e-liquids, iAsIII was detected in 59%, iAsV in 94%, and MMA in 47% of the samples. In the condensate of aerosols from vaping the e-liquids, iAsIII was detected in 100%, iAsV in 88%, and MMA in 13% of the samples. Inorganic arsenic species were predominant in e-liquids and aerosols of e-cigarettes. The concentration of iAsIII in the condensate of aerosols (median 3.27 µg/kg) was significantly higher than that in the e-liquid (median 1.08 µg/kg) samples. The concentration of inorganic arsenic in the vaping air was approximately 3.4 µg/m3, which approaches to the permissible exposure limit (10 µg/m3) set by the United States Occupational Safety and Health Administration (OSHA). According to the Environmental Protection Agency's unit risk factor (4.3 × 10-3 per µg/m3) for inhalation exposure to inorganic arsenic in the air, the estimated excess lung cancer risk from lifetime exposure to inorganic arsenic in the e-cigarette vaping air (3.4 µg/m3), assuming e-cigarette vaping at 1% of the time, is as high as 1.5 × 10-4. These results raise health concerns over the exposure to arsenic from electronic cigarettes.

Quantitative synthesis of protein-DNA conjugates with 1 : 1 stoichiometry.

We describe here a binding-facilitated reaction strategy, enabling quantitative conjugation of DNA to native proteins with a desirable 1 : 1 stoichiometry. The technique takes advantage of the iterative affinity interaction and covalent binding processes to achieve complete conjugation. The complete conjugation obviates the need for separation of the protein-DNA conjugates as required by other DNA-protein conjugation methods.

Methylated pentavalent arsenic metabolites are bifunctional inducers, as they induce cytochrome P450 1A1 and NAD(P)H:quinone oxidoreductase through AhR- and Nrf2-dependent mechanisms.

Activation of the aryl hydrocarbon receptor (AhR) ultimately leads to the induction of the carcinogen-activating enzyme cytochrome P450 1A1 (CYP1A1), and activation of the nuclear factor-erythroid 2 p45-related factor 2 (Nrf2) in addition to the AhR pathway induces the expression of the NADP(H):quinone oxidoreductase (NQO1). Therefore, the aim of this study was to examine the effect of As(III) pentavalent metabolites, MMA(V), DMA(V), and TMA(V), on AhR and Nrf2 activation and on the expression of their prototypical downstream targets CYP1A1 and NQO1, respectively. Our results showed that treatment of HepG2 cells with MMA(V), DMA(V), or TMA(V) in the absence and presence of 2,3,7,8-tetrachlorodibenzo-p-dioxin or sulforaphane significantly induced both CYP1A1 and NQO1 at the mRNA, protein, and catalytic activity levels. Furthermore, these metabolites increased the AhR-dependent XRE-driven and the Nrf2-dependent ARE-driven luciferase reporter activities, which coincided with increased nuclear accumulation of both transcription factors. However, none of these metabolites were shown to be AhR ligands. The induction of CYP1A1 by these metabolites seems to be ligand-independent, possibly through a decrease in HSP90 protein expression levels. The metabolites also increased ROS production, which was significantly higher than that produced by As(III). Upon knockdown of AhR and Nrf2 the MMA(V)-, DMA(V)-, and TMA(V)-mediated induction of both CYP1A1 and NQO1 proteins was significantly decreased. In conclusion, this study demonstrates for the first time that methylated pentavalent arsenic metabolites are bifunctional inducers, as they increase CYP1A1 by activating the AhR/XRE signaling pathway and they increase NQO1 by activating the Nrf2/ARE signaling pathway in addition to the AhR/XRE pathway.

Selection and analytical applications of aptamers binding microbial pathogens.

DNA aptamers specifically recognizing microbial cells and viruses have a range of analytical and therapeutic applications. This article describes recent advances in the development of aptamers targeting specific pathogens (e.g., live bacteria, whole viral particles, and virally-infected mammalian cells). Specific aptamers against pathogens have been used as affinity reagents to develop sandwich assays, to label and to image cells, to bind with cells for flow-cytometry analysis, and to act as probes for development of whole-cell biosensors. Future applications of aptamers to pathogens will benefit from recent advances in improved selection and new aptamers containing modified nucleotides, particularly slow off-rate modified aptamers (SOMAmers).

[Recent advances in aptamer affinity chromatography].

Aptamer affinity chromatography makes the use of aptamers as affinity ligands to achieve chromatographic separation and concentration. Aptamers are single stranded oligonucleotides that can specifically bind to targets. They show unique advantages over antibodies in preparation, stability and applications. This article summarizes the applications of aptamer affinity chromatography to the separation and analysis of small molecules, proteins, and cells. By critically evaluating literature, this review describes the current status and discusses the future prospects of aptamer affinity chromatography.