Immunoassay for pathogenic bacteria using platinum nanoparticles and a hand-held hydrogen detector as transducer. Application to the detection of Escherichia coli O157:H7.

An innovative approach is presented for portable and sensitive detection of pathogenic bacteria. A novel synthetic hybrid nanocomposite encapsulating platinum nanoparticles, as a highly efficient catalyst, catalyzes the hydrolysis of the ammonia-borane complex to generate hydrogen gas. The nanocomposites are used as a label for immunoassays. A portable hand-held hydrogen detector combined with nanocomposite-induced signal conversion was applied for point-of-care testing of pathogenic bacteria. A hand-held hydrogen detector was used as the transducer. Escherichia coli O157:H7 (E. coli O157: H7), as detection target, formed a sandwich structure with magnetic beads and hybrid nanocomposites. Magnetic beads were used for separation of the sandwich structure, and hybrid nanocomposites as catalysts to catalyze the generation of hydrogen from ammonia-borane. The generated hydrogen was detected by a hydrogen detector using an electrochemical method. E. coli O157:H7 has a detection limit of 10 CFU·mL-1. The immunosensor made the hand-held hydrogen detector a point-of-care meter to be used outdoors for the detection and quantification of targets beyond hydrogen. Graphical abstract Schematic presentation of one-pot synthetic peptide-Cu3(PO4)2 hybrid nanocomposites embedded PtNPs (PPNs), encapsulating many Pt particles. The PPNs acts as an ideal immunoprobe for hand-held H2 detector signal readouts, by transforming pathogenic bacteria recognition events into H2 signals.

Disposable syringe-based visual immunotest for pathogenic bacteria based on the catalase mimicking activity of platinum nanoparticle-concanavalin A hybrid nanoflowers.

Disposable syringes were used in a novel point-of-care visual test for detecting pathogenic bacteria (Escherichia coli O157:H7 and Salmonella typhimurium). Hybrid nanoflowers composed of platinum nanoparticles and concanavalin A (Pt-nanoflowers) were prepared through a one-pot reaction and were found to be viable catalase mimics. They catalyze the decomposition of hydrogen peroxide (H2O2) to generate O2. When used as labels in immunoassays, they integrate both the functions of biological recognition and signal amplification. The disposable syringe pressure readout was combined with Pt-nanoflower signal conversion and successfully applied to a visual bacteria detection scheme. Both Escherichia coli O157:H7 and Salmonella typhimurium can be quantified with detection limits of as low as 15 and 7 CFU·mL-1, respectively. Graphical abstract One-pot synthetic platinum nanoparticle (PtNP)-concanavalin A hybrid nanoflowers (Pt-nanoflowers), have been used as ideal signal labels for immunoassays and integrating both essential functions of biological recognition and signal amplification. Disposable syringes were used as a readout to detect pathogenic bacteria.

Hemin-incorporated nanoflowers as enzyme mimics for colorimetric detection of foodborne pathogenic bacteria.

Rapid, sensitive and point-of-care detection of foodborne pathogenic bacteria is essential for food safety. In this study, we found that hemin-concanavalin A hybrid nanoflowers (HCH nanoflowers), as solid mimic peroxidase, could catalyze oxidation of 2,2'-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt (ABTS) in the presence of H2O2 to a green-colored product. HCH nanoflowers, integrating the essential functions of both biological recognition and signal amplification, meet the requirements of signal labels for colorimetric immunoassay of bacteria. In view of the excellent peroxidase mimetic catalytic activity of HCH nanoflowers, a colorimetric biosensing platform was newly constructed and applied for sensitive detection of foodborne Escherichia coli O157:H7 (E. coli O157:H7). The corresponding detection limits was as low as 4.1 CFU/mL with wide linear ranges (101-106 CFU/mL).

Correction to: A pregnancy test strip for detection of pathogenic bacteria by using concanavalin A-human chorionic gonadotropin-Cu3(PO4)2 hybrid nanoflowers, magnetic separation, and smartphone readout.

The published version of this article, unfortunately, contained error. The authors are re-writing to express their sincere apology for a mistake that a mark "10-5, 10-4, 10-3, 10-2, 10-1 CFU•mL-1" in the legend of Fig. 2 was not corrected as "105, 104, 103, 102, 101 CFU•mL-1".

A pregnancy test strip for detection of pathogenic bacteria by using concanavalin A-human chorionic gonadotropin-Cu3(PO4)2 hybrid nanoflowers, magnetic separation, and smartphone readout.

Pregnancy test strips are widely used in daily life. A commercial pregnancy test strip was modified to obtain a point-of-care device for the detection of pathogenic bacteria. Hybrid nanoflowers were prepared from concanavalin A, human chorionic gonadotropin, and Cu3(PO4)2 via a one-pot method. They were used as signaling probes in an off-the-shelf pregnancy test strip. This modified lateral flow immunoassay can detect Escherichia coli O157:H7 with a detection limit of 4 CFU·mL-1, and Salmonella typhimurium with a detection limit of 3 CFU·mL-1. Conceivably, the method has high potential as a portable and cost-effective tool for rapid determination of a wide range of analytes, especially in resource-constrained settings. Graphical abstract Hybrid nanoflower loaded human chorionic gonadotropin (hCG) and concanavalin A (hCG - nanoflowers) were synthesized via a one-pot method and used as signal labels with commercial commercial-off-the-shelf pregnancy test strips to detect pathogenic bacteria targets, thus yielding an easily smartphone readout signal.

Modified beacon probe assisted dual signal amplification for visual detection of microRNA.

In a recent study, we reported a novel assay for the detection of microRNA-21 based on duplex-specific nuclease (DSN)-assisted isothermal cleavage and hybridization chain reaction (HCR) dual signal amplification. The Fam modified double-stranded DNA products were generated after the HCR, another biotin modified probe was digested by DSN and released from the magnetic beads after the addition of the target miRNA. The released sequence was then combined with HCR products to form a double-tagging dsDNA, which can be recognized by the lateral flow strips. In this study, we introduced a 2-OMethyl-RNA modified beacon probe (2-OMe-MB) to make some improvements based on the previous study. Firstly, the substitution of modified probe combined on magnetic beads avoids the fussy washing steps for the separation of un-reacted probes. Furthermore, the modification of 2-OMe on the stem of the probe avoided the unnecessary cleavage by DSN, which greatly reduce the background signal. Compared to the previous work, these improvements save us a lot of steps but possess the comparable sensitivity and selectivity.

Colorimetric detection of microRNA based on DNAzyme and nuclease-assisted catalytic hairpin assembly signal amplification.

Accurate and quantitative analysis of microRNA (miRNA) expression is critical for the diagnostics and theranostics of a disease. Herein, a proof-of-concept of a colorimetric horseradish peroxidase-mimicking DNAzyme (HRP-DNAzyme) biosensor for miRNA assay based on nuclease-assisted catalytic hairpin assembly (CHA) signal amplification was demonstrated. Duplex-specific nuclease (DSN) was employed to cleave the single-stranded DNA (ssDNA) chimeric probe (CP) on the magnetic bead (MB) surface via hybridization of the CP and target miRNA. The regenerated miRNA can cleave a large number of ssDNA CP to produce CHA initiator sequence fragments. The CP consists of two main regions: a target miRNA recognition DNA sequence at the 5' end and a CHA initiator (CI) sequence at the 3' end. The catalyzed assembly process of CHA produces a large amount of G-rich DNA. In the presence of hemin, the G-rich DNA forms G-quadruplex/hemin complex and mimics the horseradish peroxidase activity, which catalyzes a colorimetric reaction. For the proof-of-concept, microRNA-21 (miR-21) was selected as the model target to authenticate this strategy as a versatile assay platform. The proposed strategy allowed quantitation of the sequence specificity of miRNA-21 with a detection limit of 9.2 fM in a dynamic range from 10 fM-1 nM, with an excellent ability to discriminate the differences in miRNAs. Additionally, the miRNA assay in real samples was satisfactory, thereby confirming its applicability. Therefore, this method exhibited a great potential as a miRNA quantification method in biomedical research and clinical diagnosis.

Lateral flow nucleic acid biosensor for sensitive detection of microRNAs based on the dual amplification strategy of duplex-specific nuclease and hybridization chain reaction.

MicroRNAs (miRNAs) constitute novel biomarkers for various diseases. Accurate and quantitative analysis of miRNA expression is critical for biomedical research and clinical theranostics. In this study, a method was developed for sensitive and specific detection of miRNAs via dual signal amplification based on duplex specific nuclease (DSN) and hybridization chain reaction (HCR). A reporter probe (RP), comprising recognition sequence (3' end modified with biotin) for a target miRNA of miR-21 and capture sequence (5' end modified with Fam) for HCR product, was designed and synthesized. HCR was initiated by partial sequence of initiator probe (IP), the other part of which can hybridize with capture sequence of RP, and was assembled by hairpin probes modified with biotin (H1-bio and H2-bio). A miR-21 triggered cyclical DSN cleavage of RP, which was immobilized to a streptavidin (SA) coated magnetic bead (MB). The released Fam labeled capture sequence then hybridized with the HCR product to generate a detectable dsDNA. This polymer was then dropped on lateral flow strip and positive result was observed. The proposed method allowed quantitative sequence-specific detection of miR-21 (with a detection limit of 2.1 fM, S/N = 3) in a dynamic range from 100 fM to 100 pM, with an excellent ability to discriminate differences in miRNAs. The method showed acceptable testing recoveries for the determination of miRNAs in serum.

Visual detection of nucleic acids based on lateral flow biosensor and hybridization chain reaction amplification.

In this study, a new lateral flow nucleic acid biosensor (LFNAB) using hybridization chain reaction (HCR) for signal amplification was developed for visual detection of nucleic acids with high sensitivity and low cost. A "sandwich-type" detection strategy was employed in our design. The sandwich system of capture probe (CP)/target DNA/reporter probe (RP)-HCR complexes was fabricated as the sensing platform. As the initiator strand, reporter probe propagated a chain reaction of hybridization events between the two hairpin probes modified with biotin, and determined whether long nicked DNA polymers were formed. The biotin-labeled double-strand DNA polymers then introduced numerous Streptavidin (SA)-labeled gold nanoparticles (AuNPs) on the lateral flow device. The CP/target DNA/RP-HCR complexes were captured on the test zone by the specific reaction between anti-Fam monoclonal antibody (anti-Fam mAb) on the test zone and Fam of the complexes. The accumulation of AuNPs on the test zone of the biosensor enabled the visual detection of specific sequences. The detection limit of specific DNA was as low as 1.76pM, which was about 2 orders lower than that of the LFNAB without HCR amplification. And the detection limit of Salmonella was 3×103cfumL-1. In conclusion, this visual detection system, HCR-LFNAB, is suitable for non-specialist personnel and point-of-care (POC) diagnosis in low-resource settings.

The Impact of Transformational Leadership on Safety Climate and Individual Safety Behavior on Construction Sites.

Unsafe acts contribute dominantly to construction accidents, and increasing safety behavior is essential to reduce accidents. Previous research conceptualized safety behavior as an interaction between proximal individual differences (safety knowledge and safety motivation) and distal contextual factors (leadership and safety climate). However, relatively little empirical research has examined this conceptualization in the construction sector. Given the cultural background of the sample, this study makes a slight modification to the conceptualization and views transformational leadership as an antecedent of safety climate. Accordingly, this study establishes a multiple mediator model showing the mechanisms through which transformational leadership translates into safety behavior. The multiple mediator model is estimated by the structural equation modeling (SEM) technique, using individual questionnaire responses from a random sample of construction personnel based in Hong Kong. As hypothesized, transformational leadership has a significant impact on safety climate which is mediated by safety-specific leader-member exchange (LMX), and safety climate in turn impacts safety behavior through safety knowledge. The results suggest that future safety climate interventions should be more effective if supervisors exhibit transformational leadership, encourage construction personnel to voice safety concerns without fear of retaliation, and repeatedly remind them about safety on the job.

Unchanged survival rates of Shadoo knockout mice after infection with mouse-adapted scrapie.

Previous studies have demonstrated that Shadoo (Sho), a GPI-linked glycoprotein encoded by the Sprn gene with a membrane localization similar to PrP(C), is reduced in the brains of rodents with terminal prion disease. To determine the functional significance of Sho in prion disease pathogenesis, Sho-deficient mice were generated by gene targeting. Sho knockout and control wild-type (WT) mice were infected with themouse-adapted scrapie strains 22L or RML. No significant differences in survival, the incubation period of prion disease or other disease features were observed between Sho mutant and WT mice. In this model of prion disease, Sho removal had no effect on disease pathogenesis.

ABCG5/8 variants are associated with susceptibility to coronary heart disease.

ATP-binding cassette sub-family G member 5 (ABCG5) and ABCG8 are members of an ATP-binding cassette transporter superfamily. ABCG5 and ABCG8 variants affected serum levels of cholesterol and were considered as risk factors for coronary heart disease (CHD). The present control study analyzed ABCG5 and ABCG8 variants in a population for association with the risk of CHD. A total of 417 CHD patients and 267 controls were recruited for genotyping of four single nucleotide polymorphisms (SNPs; i.e. i7892T>C in ABCG5 and Tyr54CysA>G, Thr400LysC>A and 5U145A>C in ABCG8) using quantitative PCR high-resolution melting (qPCR-HRM). Serum lipid levels were measured using an automatic biochemical analyzer. The association of ABCG5/8 variants with lipid levels was analyzed using a Chi-square test. The impact of candidate ABCG5/8 SNPs on CHD was evaluated in a dominant genetic model with stepwise multiple regression analysis. Subgroup analyses were performed with regard to these SNPs, tobacco smoking status, alcohol consumption and gender. Genotypic and allelic frequencies of ABCG8 Thr400LysC>A were significantly different (P<0.05) between CHD patients and controls. CC homozygotes of the ABCG8 Thr400LysC>A SNP had greater triglyceride levels than CA/AA carriers with CHD. Logistic analysis revealed CHD risk was significantly higher in CC homozygotes of ABCG8 Thr400LysC>A than in carriers of the A allele (adjusted P=0.048; OR=2.034; 95% CI=0.983-4.207). Furthermore, there was a significant gene-tobacco smoking interaction. CC homozygotes of ABCG8 Thr400LysC>A SNP had significantly higher triglyceride concentrations (P=0.012) and an increased risk of CHD than tobacco smoking carriers of the A allele. The data from the current study suggested that ABCG8 Thr400LysC>A SNP genetic variants modulated plasma triglyceride levels and thereby affected CHD risk in the population studied. The genetic variant of ABCG8 also contributed to CHD risk through interaction with tobacco smoking.