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An ultrasensitive photoelectrochemical (PEC) bioassay for determination of microRNA was proposed based on an integrative photoactive heterojunction nanomaterial to provide the basis of excellent PEC responses and an efficient redox cycling amplification system to improve the detection performances. To establish the bioassay system, the biosensor was firstly modified with Bi2WO6@Bi2S3 and alkaline phosphatase (ALP). The detection solution was composed of ascorbic acid phosphate (AAP) and ferrocenecarboxylic acid (FcA), where ALP converted AAP into ascorbic acid (AA) to trigger a process of redox cycling amplification by reducing FcA+ to FcA, resulting in enhanced photocurrent responses of Bi2WO6@Bi2S3. In the presence of microRNA 21, it could trigger a hybridization chain reaction via the special designed hairpin DNA to produce a long repeated DNA sequences to inhibit ALP activity. Thus the reduced ALP activity and consequently decreased photocurrent signal could be obtained for detection of microRNA 21. As expected, this bioassay system performed the satisfactory performances for the ultrasensitive detection of microRNA 21 in the range from 1 fM to 1 nM with an experimental detection limit of 0.26 fM and acceptable practical applicability. Collectively, an efficient PEC bioassay for microRNA 21 is established and this strategy can be expanded to detect other microRNAs, even other molecules in cells.

Resistance to 5-fluorouracil (5-FU) and its induced immune suppression have prevented its extensive application in the clinical treatment of breast cancer. In this study, the combined effect of 50 Hz-EMFs and 5-FU in the treatment of breast cancer was explored. MCF-7 and MCF10A cells were pre-exposed to 50 Hz-EMFs for 0, 2, 4, 8 and 12 h and then treated with different concentrations of 5-FU for 24 h; cell viability was analyzed by MTT assay and flow cytometry. After pre-exposure to 50 Hz-EMFs for 12 h, apoptosis and cell cycle distribution in MCF-7 and MCF10A cells were detected via flow cytometry and DNA synthesis was measured by EdU incorporation assay. Apoptosis-related and cell cycle-related gene and protein expression levels were monitored by qPCR and western blotting. Pre-exposure to 50 Hz-EMFs for 12 h enhanced the antiproliferative effect of 5-FU in breast cancer cell line MCF-7 in a dose-dependent manner but not in normal human breast epithelial cell line MCF10A. Exposure to 50 Hz-EMFs had no effect on apoptosis and P53 expression of MCF-7 and MCF10A cells, whereas it promoted DNA synthesis, induced entry of MCF-7 cells into the S phase of cell cycle, and upregulated the expression levels of cell cycle-related proteins Cyclin D1 and Cyclin E. Considering the pharmacological mechanisms of 5-FU in specifically disrupting DNA synthesis, this enhanced inhibitory effect might have resulted from the specific sensitivity of MCF7 cells in active S phase to 5-FU. Our findings demonstrate the enhanced cytotoxic activity of 5-FU on MCF7 cells through promoting entry into the S phase of the cell cycle via exposure to 50 Hz-EMFs, which provides a novel method of cancer treatment based on the combinatorial use of 50 Hz-EMFs and chemotherapy.

Conductive carbon black (CB) nano-particle filled silicone elastomer (SE) composites with dimethyl silicone oil (SO) as a diluting agent and plasticizer were fabricated for potential strain sensing applications. Heat treatment was used to stabilize the composites, and thermal gravimetric analysis confirmed the good stability of the composites after heat treatment. Effect of CB loading on electrical properties of the composites was studied and a percolation transition range of 0.5–2.5 wt% was observed. I–V characteristics and impedance analysis were used to reveal the conductive mechanisms of the composites. The presence of SO reduced the Young's modulus of the composites without lowering the elongation at break. Characterization of coupled electromechanical properties of the composites demonstrates that, in the post-percolation range, the 9.0CB composite possesses suitable strain sensitivity, good repeatability and linearity as well as slight strain rate dependence, so that it can be used in flexible strain sensors for measuring repeated large deformations.

Electrochemical immunosensors have attracted great interest in the search for a selective, simple and reliable system for molecular recognition. Presently, electrochemical immunosensors have been widely studied for biomedical molecular's detection, but the regeneration of these immunosensors has restricted their wide application. To prepare a regeneration-free immunosensor, which may be more suitable for clinical determination, a repeatable immunoassay system was developed based on an electrochemical immunosensor with magnetic nanoparticles, biotin-avidin system (BAS) and Fab antibodies for the heart failure markers aminoterminal pro-brain natriuretic peptides (NT-proBNP). At the same time, a microfluidic system was combined into the proposed system, which enabled continuous determination. Using NT-proBNP as a model system, the proposed immunosensor exhibited rapid and sensitive amperometric response to NT-proBNP with good selectivity, stability, and a wide linear range (0.005-1.67 ng/mL and 1.67-4 ng/mL with a detection limit of 0.003 ng/mL under optimal conditions). Importantly, the proposed immunosensor was also suitable for the detection of other proteins and provided new opportunities for disease diagnosis.

N-Terminal pro-brain natriuretic peptide (NT-proBNP) is an important marker for heart failure that reflects ventricular volume expansion, ventricular overload and the degree of cardiac injury. To establish a sensitive and simple detection method for it in serum, a regeneration-free immunosensor with novel Fab fragment monoclonal antibodies was prepared. The sensor detected NT-proBNP from 0.04 to 2.5 ng/ml with a limit of 0.03 ng/ml. The immunosensor is therefore a simple, cost-effective method to detect NT-proBNP and the proposed immunoassay system would enable other proteins to be detected and open new opportunities for protein diagnostics.

A high activity creatine amidinohydrolase (creatinase) from Arthrobacter nicotianae 02181 (a strain newly isolated from soil which may utilize creatinine as the unique organic source) was purified, characterized and the creatinase gene was cloned and analyzed in this study. Cells were cultivated under optimized condition for enzyme yield and creatinase was purified by the DEAE-cellulose and hydroxylapatite (HA) chromatography. The creatinase was found to be a dimmer formed by two identical subunit of 46.4 kDa, and the specific activity of the purified creatinase reached 124.44 U/mg protein, which was about 13 folds of the maximum value ever reported. The enzyme was found to be most active at 37 °C (pH 7.0), and it was found to be relatively stable bellow 45 °C around pH 7.0 by fluorescence spectroscopy and circular dichroism (CD) analysis. The activity of this creatinase could be significantly inhibited by Cu2+, Hg2+, Fe3+and SDS, and it could be improved by Ca2+ and NaN3.The creatinase gene was cloned by the consensus-degenerate hybrid oligonucleotide primers (CODEHOP) PCR and the genome walking method. Nucleotide sequence analysis of this gene revealed an open reading frame (ORF) of 1254 base pair (bp) encoding a 417 amino acid (aa) protein. The primary amino acid sequence alignment search in the database revealed a moderate homology between the deduced amino acid sequence and other creatinase. The sequence has been submitted to Genbank with the accession number EU004199 .

A novel electrochemical immunosensor for sensitive detection of cardiac biomarker N-terminal pro-B-type natriuretic peptide (NT-proBNP) is fabricated based on the nanostructural gold and carbon nanotubes composite as desirable platform for the capture antibodies immobilization and gold nanochains (AuNCs) and horseradish peroxidase (HRP) complex labeled secondary antibodies (AuNCs-HRP-Ab(2)) for signal amplification. The gold nanochains were prepared by the employment of L-ascorbic acid (AA) as a mediator and template. With the surface area enhancement by nanostructural gold functionalized carbon nanotubes composite, the amount of immobilized primary antibodies (Ab(1)) can be enhanced. More importantly, enhanced sensitivity can be achieved by introducing the multibioconjugates of AuNCs-HRP-Ab(2) onto the electrode surface through "sandwich" immunoreactions. The linear range extended from 0.02 to 100 ng/mL with a correlation coefficient of R=0.997 and a limit of detection reaching 6 pg/mL at a signal-to-noise ratio of 3:1, which is well below the commonly accepted concentration threshold (0.1 ng/mL) used in clinical diagnosis. The specificity, regeneration and stability test demonstrated the feasibility of the developed immunoassay, which gives the attractive characteristics to be a candidate for the detection of NT-proBNP and other proteins of interest in both fundamental and applied research.