OBJECTIVETo develop a quartz crystal microbalance (QCM) immunosensor with high sensitivity and selectivity for the rapid detection of diethylstilbestrol.

METHODSDextran was used as reducing agent for preparing gold nanoparticles (AuNPs) with the size of 40 nm. The AuNPs were coupled with anti-DES antibody after amination. A monolayer was generated after immersing the quartz crystal into the solution of 5 mmol/L 11-mercaptoundecanoic acid(MUA) for 16 hours. After the monolayer was activated by 1-ethyl-3-(3-dimethylaminopropry) carbodiimide hydrochloride (EDC·HCl) and N-hydrosuccinimide (NHS), 20 μl of 2.2 mg/ml DES-HS-BSA was dropped onto the surface of crystal to prepare a sensitive membrane which can recognize DES specifically. Then, 50 μl of 1 mol/L ethanolamine (pH 8.5) was used to seal the carboxylic groups to make the sensitive membrane which could identify DES specifically. QCM immunosensor was used as detection platform to optimize the reaction conditions. Under the optimized conditions, 10 μl of 28 μg/ml AuNPs-antibody was mixed with 10 μl of 0.03-2.5 μg/ml DES, and the mixture was added on the sensitive membrane. QCM immunosensor was used to detect the signals and the standard curve was obtained at the same time. The detection limit was calculated based on the standard curve. The specificity was evaluated by testing DES and its analogues with the same concentration.

RESULTSThe optimized concentration for the immobilization of DES-HS-BSA on the surface of QCM was 2.2 mg/ml. The optimized concentration for coupling anti-DES antibody with AuNPs was 7 μg/ml and 15 nmol/L, respectively. The optimized concentration of AuNPs-antibody was 14 μg/ml. The logarithm of DES concentration was proportional to the frequency shift in the range of 0.16-500 ng/ml, Δf=-24.170 lgCDES+69.71, R(2)=0.998. The detection limit of this method was 0.13 ng/ml. DES analogues could not influence the detection of DES obviously, so the sensor had good specificity.

CONCLUSIONThe quartz crystal microbalance immunosensor with gold nanoparticals amplification could detect DES sensitively and rapidly.

Biosensing Techniques ; Diethylstilbestrol ; isolation & purification ; Gold ; Limit of Detection ; Nanoparticles ; Quartz Crystal Microbalance Techniques

Objective To characterize the urinary microbial profile of male non-muscle invasive bladder cancer patients compared to healthy controls.Methods Between March,2017 and September,2017,mid-stream urine from 26 non-muscle invasive bladder cancer and 18 non-neoplastic controls were collected by the clean method,then centrifuged and processed for 16S rRNA gene sequencing.Sequencing reads were processed for evaluating alpha diversity and beta diversity using QIIME.LEfSe algorithm was performed to identify potential bacterial genera biomarker.Results The smoking cases were more in tumor group than those in control group(21 vs.7,P ＜ 0.01).The urinary microenvironment of bladder cancer was characterized by increased bacterial richness (Observed species index,Chaol index and Ace index,125.77 ± 69.64 vs.80.38 ± 46.24;142.82 ± 76.74 vs.90.68 ± 47.62;and 147.92 ± 77.68 vs.88.19 ± 45.38,all P ＜ 0.05) and by the enrichment of some bacterial genera (e.g.,Acinetobacter and Anaerococcus).Significant difference in β diversity was found between cancer and non-cancer group (ANOSIM,P =O.009).A clear hierarchical clustering of cancer samples was observed,suggesting a common dysbiosis associated to bladder cancer.Conclusions Patients with non-muscle invasive bladder cancer exhibit a different microbial community compared to non-neoplastic controls,suggesting a possible pathophysiological correlation between microbiome and bladder cancer.Urinary microbial community may be associated with the prognosis of NMIBC.

The study aims to explore the effect of mesenchymal stem cells-derived exosomes (MSCs-Exo) on staurosporine (STS)-induced chondrocyte apoptosis before and after exposure to pulsed electromagnetic field (PEMF) at different frequencies. The AMSCs were extracted from the epididymal fat of healthy rats before and after exposure to the PEMF at 1 mT amplitude and a frequency of 15, 45, and 75 Hz, respectively, in an incubator. MSCs-Exo was extracted and identified. Exosomes were labeled with DiO fluorescent dye, and then co-cultured with STS-induced chondrocytes for 24 h. Cellular uptake of MSC-Exo, apoptosis, and the protein and mRNA expression of aggrecan, caspase-3 and collagenⅡA in chondrocytes were observed. The study demonstrated that the exposure of 75 Hz PEMF was superior to 15 and 45 Hz PEMF in enhancing the effect of exosomes in alleviating chondrocyte apoptosis and promoting cell matrix synthesis. This study lays a foundation for the regulatory mechanism of PEMF stimulation on MSCs-Exo in inhibiting chondrocyte apoptosis, and opens up a new direction for the prevention and treatment of osteoarthritis.
Animals ; Rats ; Apoptosis ; Chondrocytes ; Electromagnetic Fields ; Exosomes/physiology* ; Mesenchymal Stem Cells/metabolism*