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A novel electrochemical immunosensor based on the integration of immunomagnetic separation and bienzymatic amplification for sensitive detection of virus particles was fabricated in this work. The bienzymatic strategy was realized by using the first enzyme as tracer tagged on immunomagnetic beads which could be accumulated on the magneto controlled home-made Au electrode (m-AuE) and the second enzyme immobilized on the m-AuE by layer-by-layer (LBL) assembly technique. The proposed immunosensor not only provides a rapid, simple, cost-effective and on-site platform with high sensitivity, selectivity, and reproducibility for early diagnosis but also presents a new approach for sensitive magneto immunoassay. Keywords: Virus detection, Immunosensor, Magnetic bead, Electrochemical detection, Bienzymatic strategy, Layer-by-layer assembly

A sandwich immunoassay method for rapid detection of dual cancer biomarkers in serum on a magnetic field controllable microfluidic chip (MFCM-Chip) was established. A nickel pattern was used to generate high magnetic field gradients to increase the magnetic force on the superparamagnetic beads (SPMBs), which enabled the rapid generation of controllable SPMB patterns in microfluidic channels. The SPMB patterns could keep stable during the fast continuous washing process even at a flow rate of 50 μL/min. This approach demonstrated excellent specificity because the fast continuous washing could remove non-specifically adsorptive contaminants more efficiently than fixed volume batch washing. This approach was used to simultaneously detect carcinoma embryonic antigen (CEA) and α-fetoprotein (AFP) directly in serums. The whole on-chip detection was finished within 40 min, which was much faster than conventional enzyme-linked immunosorbent assay (ELISA) method. High luminescent streptavidin modified QDs (SA-QDs) were used as fluorescence indicators, and the detection limits were 3.5 ng/mL and 3.9 ng/mL for CEA and AFP, respectively. The linear ranges were from 10.0 ng/mL to 800.0 ng/mL. With the high sensitivity, high selectivity and short assay time, this approach could be used for rapid, high throughput detection of cancer biomarkers in clinical trials.

To evaluate factors in penile squamous cell carcinoma predictive of pelvic lymph node metastasis and survival. Data were collected and analyzed retrospectively in 146 patients with squamous cell carcinoma of penis who underwent bilateral inguinal lymph node dissection in our center between January 1998 and April 2011. Variables recorded included serum squamous cell carcinoma antigen, primary tumor p53 immunoreactivity, histological grade, pathological tumor stage, lymphatic or vascular invasion, absent/unilateral or bilateral inguinal lymph node involvement, number of metastatic inguinal lymph nodes, presence of extracapsular growth and lymph node density. Seventy patients had inguinal lymph node metastasis (LNM). Of these, 33 (47.1 %) had pelvic LNM. Primary tumor strong p53 expression, lymphatic or vascular invasion, involvement of more than two inguinal lymph nodes and 30 % or greater lymph node density were significant predictors of pelvic LNM. Primary tumor strong p53 expression (odds ratio [OR] 5.997, 95 % confidence intervals [CI] 1.615–22.275), presence of extracapsular growth (OR 2.209, 95 % CI 1.166–4.184), involvement of more than two inguinal lymph nodes (OR 2.494, 95 % CI 1.086–5.728) and pelvic lymph node involvement (OR 18.206, 95 % CI 6.807–48.696) were independent prognostic factors for overall survival. Primary tumor expression of p53, lymphatic or vascular invasion, number of metastatic inguinal lymph nodes and lymph node density were all predictors of pathologic pelvic lymph node involvement. Patients with pelvic LNM had an adverse prognosis, with a 3-year overall survival rate of approximately 12.1 %. Pelvic lymph node dissection should be considered in these cases.

Quantum dots (QDs) with fluorescence in the second near-infrared window (NIR-II, 1000-1400 nm) are ideal fluorophores for in vivo imaging of deep tissue with high signal-to-noise ratios. Ag₂Se (bulk band gap 0.15 eV) is a promising candidate for preparing NIR-II QDs. By using 1-octanethiol as ligand to effectively balance the nucleation and growth, tuning the fluorescence of Ag₂Se QDs was successfully realized in the NIR-II window ranged from 1080 to 1330 nm. The prepared Ag₂Se QDs can be conveniently transferred to the aqueous phase by ligand exchange, showing great potential for multicolor NIR-II fluorescence imaging in vivo.

Many viruses invade host cells by entering the cells and releasing their genome for replication, which are remarkable incidents for viral infection. Therefore, the viral internal and external components should be simultaneously labeled and dynamically tracked at single-virus level for further understanding viral infection mechanisms. However, most of the previously reported methods have very low labeling efficiency and require considerable time and effort, which is laborious and inconvenient for researchers. In this work, we report a general strategy to high-efficiently label viral envelope and genome for single-virus imaging with quantum dots (QDs) and Syto 82, respectively. It was found that nearly all viral envelopes could be labeled with QDs with superior stability, which makes it possible to realize global and long-term tracking of single virus in individual cells. Effectively labeling their genome with Syto 82, about 90% of QDs-labeled viruses could be used to monitor the viral genome signal, which may provide valuable information for deeply studying viral genome transport. This is very important and meaningful to investigate the viral infection mechanism. Our labeling strategy has advantage in commonality, convenience and efficiency, which is expected to be widely used in biological research.

Single molecule electrochemistry (SME) has gained much progress in fundamental studies, but it is difficult to use in practice due to its less reliability. We have solved the reliability of single molecule electrochemical detection by integration of digital analysis with efficient signal amplification of enzyme-induced metallization (EIM) together with high-throughput parallelism of microelectrode array (MA), establishing a digital single molecule electrochemical detection method (dSMED). Our dSMED has been successfully used for alkaline phosphatase (ALP) detection in the complex sample of liver cancer cells. Compared to direct measurement of the oxidation current of enzyme products, EIM can enhance signals by about 100 times, achieving signal-to-background ratio high enough for single molecule detection. The integration of digital analysis with SME can further decrease the detection limit of ALP to 1 aM relative to original 50 aM, enabling dSMED to be sensitively, specifically and reliably applied in liver cancer cells. The presented dSMED is enormously promising in exploring physical and chemical properties of single molecules, single biomolecular detection, or single-cell analysis.

Direct analysis of biomolecules in complex biological samples remains a major challenge for fluorescence-based approaches due to the interference of background signals. Herein, we report an analytical methodology by exploiting a single low-cost near-infrared sub-nanosecond pulse laser to synchronously actualize optical trapping and two-photon excitation fluorescence for senstive detection of carcinoembryonic antigen (CEA) in buffer solution and human whole serum with no separation steps. The assay is performed by simultaneously trapping and exciting the same immune-conjugated microsphere fabricated with a sandwich immunization strategy. Since the signal is strictly limited in the region of a three-dimensional focal volume where the microsphere is trapped, no obvious background signal is found to contribute the detected signals and thus high signal-to-background data are obtained. As a proof-of-concept study, the constructed platform exhibits good specificity for CEA and the detection limit reaches as low as 8pg/mL (45 fM) with a wide linear range from 0.01 to 60ng/mL in the both cases. To investigate the potential application of this platform in clinical diagnosis, 15 cases of serum samples were analyzed with satisfactory results, which further confirm the applicability of this method.

Theranostic nanoprobes integrated with diagnostic imaging and therapy capabilities have shown great potential for highly effective tumor therapy by realizing imaging-guided drug delivery and tumor treatment. Developing novel high-performance nanoprobes is an important basis for tumor theranostic application. Here, near-infrared (NIR) fluorescent and low-biotoxicity Ag

Due to excellent optical properties, quantum dots (QDs) have been widely applied to sensing, labeling, and imaging. For the fabrication of QD-based bioprobes, purification is usually the crucial step. Hydrophilic octylamine grafted polyacrylic acid modified QDs (OPA-QDs) were prepared, and purified by high-performance size exclusion chromatography (HPSEC) to remove excess OPA and aggregated QDs. The percentage of suspended agglomerates of OPA-QDs in the unpurified OPA-QDs increases from 4% to 31% through a year, but the purified OPA-QDs of the same batch possess excellent colloidal stability for at least one year. Subsequently, QD-based bioprobes were fabricated by the conjugation between QDs and streptavidin (SA) or antibody (IgG), generating QD-SA and QD-IgG, respectively, which were purified via HPSEC. Finally, the resulting QD-SA and QD-IgG were adopted to detect tumour markers on slices and showed specific positive signals without nonspecific adsorption, which was contrary to the unpurified QD-IgG. Thus, the HPSEC-coupled system proposed in the current work is potent and universal for the generation of purified and monodisperse QD-based bioprobes, which is promising in the nanobiodetection field.

Cell-derived microvesicles (MVs) are natural carriers that can transport biological molecules between cells, which are expected to be promising delivery vehicles for therapeutic purposes. Strategies to label MVs are very important for investigation and application of MVs. Herein, ultrasmall Mn-magnetofunctionalized Ag2Se quantum dots (Ag2Se@Mn QDs) integrated with excellent near-infrared (NIR) fluorescence and magnetic resonance (MR) imaging capabilities have been developed for instant efficient labeling of MVs for their in vivo high-resolution dual-mode tracking. The Ag2Se@Mn QDs were fabricated by controlling the reaction of Mn(2+) with the Ag2Se nanocrystals having been pretreated in 80 °C NaOH solution, with an ultrasmall size of ca. 1.8 nm, water dispersibility, high NIR fluorescence quantum yield of 13.2%, and high longitudinal relaxivity of 12.87 mM(-1) s(-1) (almost four times that of the commercial contrast agent Gd-DTPA). The ultrasmall size of the Ag2Se@Mn QDs enables them to be directly and efficiently loaded into MVs by electroporation, instantly and reliably conferring both NIR fluorescence and MR traceability on MVs. Our method for labeling MVs of different origins is universal and free of unfavorable influence on intrinsic behaviors of MVs. The complementary imaging capabilities of the Ag2Se@Mn QDs have made the long-term noninvasive whole-body high-resolution dual-mode tracking of MVs in vivo realized, by which the dynamic biodistribution of MVs has been revealed in a real-time and in situ quantitative manner. This work not only opens a new window for labeling with QDs, but also facilitates greatly the investigation and application of MVs.

Real-time, long-term, single-particle tracking (SPT) provides us an opportunity to explore the fate of individual viruses toward understanding the mechanisms underlying virus infection, which in turn could lead to the development of therapeutics against viral diseases. However, the research focusing on the virus assembly and egress by SPT remains a challenge because established labeling strategies could neither specifically label progeny viruses nor make them distinguishable from the parental viruses. Herein, we have established a temporally controllable capsid-specific HaloTag labeling strategy based on reverse genetic technology. VP26, the smallest pseudorabies virus (PrV) capsid protein, was fused with HaloTag protein and labeled with the HaloTag ligand during virus replication. The labeled replication-competent recombinant PrV harvested from medium can be applied directly in SPT experiments without further modification. Thus, virus infectivity, which is critical for the visualization and analysis of viral motion, is retained to the largest extent. Moreover, progeny viruses can be distinguished from parental viruses using diverse HaloTag ligands. Consequently, the entire course of virus infection and replication can be visualized continuously, including virus attachment and capsid entry, transportation of capsids to the nucleus along microtubules, docking of capsids on the nucleus, endonuclear assembly of progeny capsids, and the egress of progeny viruses. In combination with SPT, the established strategy represents a versatile means to reveal the mechanisms and dynamic global picture of the life cycle of a virus.

With unique and tunable photoluminescent properties, carbon nanodots (CNDs), as a new class of optical tags, have been extensively studied. In this chapter, we introduce the basic knowledge with respect to CNDs, including their structures and compositions, optical properties and applications in the bioimaging and biosensors. In particular, the photoluminescence (PL) mechanisms of CNDs, which are able to instructively improve its optical properties, have been emphasized and discussed in details. We hope to inspire research into the origins of the unique properties of CNDs and intrigue the researchers with different research backgrounds to participate in this field and explore the PL mechanisms of CNDs.

Tumor microenvironment has been increasingly recognized as a complex and dynamic cancer society influencing tumor invasion and progression. The prognostic significance of this microenvironment is yet to be fully appreciated. A holistic approach to obtaining integrated information on key components in tumor microenvironment is essential. Here we reported on a quantum dots (QDs)-based simultaneous in-situ detection of infiltrating macrophages, tumor microvessels density (MVD) and neovessels maturity, in gastric cancer tissues, to obtain integrated information on these components, termed as combined tumor stromal features. These stromal features had the comparable prognostic value for overall survival, and even better prognostic value for disease-free survival, compared with traditional tumor cell-based clinico-pathological parameters. Subgroups of gastric cancer patients with favorable and unfavorable combined tumor stromal features were identified, with significantly different clinical outcomes. This study demonstrated the technical advantages of QDs-based simultaneous detection of multiple biomarkers in situ, revealed the important role of tumor stroma in cancer biology, and opened a new field to predict clinical outcome in gastric cancer from the perspectives of tumor microenvironment.

BACKGROUND. The objective of this study was to evaluate the effect of pericyte coverage (PC) of differentiated tumor microvessels on the prognosis of patients with clear cell renal cell carcinoma (CCRCC). METHODS. Samples from 2 cohorts of patients with CCRCC (101 Asian patients and 524 US patients) were prepared using 2 different histologic approaches: routine sectioning versus tissue microarray. Then, the samples were immunohistochemically doubled-stained for a pericyte marker (alpha smooth muscle actin [α-SMA]) and a differentiated vessel marker (cluster of differentiation 34 [CD34]), followed by multispectral image capturing and computerized image analyses to quantify the microvessel density (MVD) and the PC of differentiated vessels. The correlations of PC and the MVD:PC ratio with clinicopathologic characteristics were analyzed. RESULTS. There was an inverse correlation between differentiated MVD and PC. Higher PC correlated with more aggressive clinicopathologic characteristics of CCRCC in both cohorts, including more advanced T-classification, higher pathologic grades, and the occurrence of tumor necrosis. The MVD:PC ratio was an independent favorable prognostic factor for overall and recurrence-free survival in the Asian cohort and for recurrence-free survival in the US cohort. PC also was an independent prognostic factor, with higher PC predicting a poorer outcome. The combination of PC and MVD was better at distinguishing the outcome of patients with CCRCC. PC combined with differentiated MVD or with the MVD:PC ratio provided additional, independent prognostic information to the Leibovich risk model, and that information was used to generate improved risk models. CONCLUSIONS. The authors consistently observed that higher PC was correlated with more aggressive clinicopathologic characteristics. PC was an independent unfavorable prognostic factor. The authors concluded that pericytes should be considered for therapeutic targeting. Cancer 2013. © 2012 American Cancer Society.

Objective The aim of this study was to evaluate and compare the measurement accuracy of the temporomandibular joint (TMJ) space in the Promax 3D cone-beam computerized tomography (CBCT) images scanned with 2 different dental protocols. Study Design TMJ space impression models were made according to the occlusion. Forty joints were scanned with the standard and the large view protocol of the Promax 3D CBCT scanner. Two observers measured the joint spaces 3 times on both radiographs and the photocopies of the impression models. Results A total of 120 CBCT images were measured. There were no significant differences among the actual joint spaces and the CBCT measurements performed with the 2 scanning protocols ( P = .305). The inter- and intraobserver variabilities were not significant. Conclusions The 2 scanning protocols provided by the Promax 3D CBCT scanner were reliable and similar for recording the TMJ space.

In this work, robust approach for a highly sensitive point-of-care virus detection was established based on immunomagnetic nanobeads and fluorescent quantum dots (QDs). Taking advantage of immunomagnetic nanobeads functionalized with the monoclonal antibody (mAb) to the surface protein hemagglutinin (HA) of avian influenza virus (AIV) H9N2 subtype, H9N2 viruses were efficiently captured through antibody affinity binding, without pretreatment of samples. The capture kinetics could be fitted well with a first-order bimolecular reaction with a high capturing rate constant k(f) of 4.25 × 10(9) (mol/L)(-1) s(-1), which suggested that the viruses could be quickly captured by the well-dispersed and comparable-size immunomagnetic nanobeads. In order to improve the sensitivity, high-luminance QDs conjugated with streptavidin (QDs-SA) were introduced to this assay through the high affinity biotin-streptavidin system by using the biotinylated mAb in an immuno sandwich mode. We ensured the selective binding of QDs-SA to the available biotin-sites on biotinylated mAb and optimized the conditions to reduce the nonspecific adsorption of QDs-SA to get a limit of detection low up to 60 copies of viruses in 200 μL. This approach is robust for application at the point-of-care due to its very good specificity, precision, and reproducibility with an intra-assay variability of 1.35% and an interassay variability of 3.0%, as well as its high selectivity also demonstrated by analysis of synthetic biological samples with mashed tissues and feces. Moreover, this method has been validated through a double-blind trial with 30 throat swab samples with a coincidence of 96.7% with the expected results.

Labeling of virus opens new pathways for the understanding of viruses themselves and facilitates the utilization of viruses in modern biology, medicine, and materials. Based on the characteristic that viruses hijack their host cellular machineries to survive and reproduce themselves, a host-cell-assisted strategy is proposed to label enveloped viruses. By simply feeding Vero cells with commercial 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(cap biotinyl) (sodium salt) (Biotin-Cap-PE), we obtained biotinylated Vero cells whose membrane systems were modified with biotin. Subsequently, pseudorabies viruses (PrV) were cultivated in the biotinylated Vero cells, and the PrV progenies were spontaneously labeled with Biotin-Cap-PE during viral natural assembly process. Since the viral natural assembly process was employed for the labeling, potential threats of genetic engineering and difficulties in keeping viral natural bioactivity were avoided. Importantly, this labeling strategy for enveloped virus greatly reduces the technical complexity and allows researchers from different backgrounds to apply it for their specified demands.

Fluorescent nanoparticles have attracted much attention over the last two decades. Due to the size- and composition-dependent optical and electrical properties, fluorescent nanoparticles have been emphasized in electronic, optical and biomedical applications. Till now, many kinds of methods have been developed to fabricate diverse fluorescent nanoparticles, which include pyrolysis, template synthesis, hydrothermal synthesis, microemulsion, electrochemical methods and so on. Among them, electrochemical methods are favored for relatively good controllability, ease of operation and mild reaction conditions. By adjusting the applied potential, current, components of the electrolyte and other relevant parameters, the fluorescent nanoparticles could be electrochemically manufactured with tunable sizes, compositions and surface structure, which allows for the modification of electronic and optical properties. Therefore, electrochemical methods are regarded as important means in preparing fluorescent nanoparticles. This review focuses on the recent progress in electrochemical fabrications of fluorescent nanoparticles (together with their optical properties and some applications in optoelectronics and biomedicine).

Ultrabright carbon nanodots-hybridized silica nanospheres (CSNs) are synthesized through the Stober process of silane functionalized C-dots. The fluorescence of carbon nanodots is converged intensely. A CSN is about 3800 times brighter than a single-carbon nanodot. Along with their high brightness and low cytotoxicity, CSNs also indicate their potential application in cellular labeling.

Real-time tracking of fluorophore-tagged viruses in living cells can help uncover virus infection mechanisms. Certainly, the indispensable prerequisite for virus-tracking is to label viruses with some bright and photostable beacons such as quantum dots (QDs) via an appropriate labeling strategy. Herein, we devise a convenient hydrazine-aldehyde based strategy to label viruses with QDs through the conjugation of 4-formylbenzoate (4FB) modified QDs to 6-hydrazinonicotinate acetone hydrazone (HyNic) modified viruses under mild conditions. On the basis of this strategy, viruses can be successfully labeled with QDs with high selectivity, stable conjugation, good reproducibility, high labeling efficiency of 92-93% and maximum retention of both fluorescence properties of QDs and infectivity of viruses, which is very meaningful to tracking and statistical analysis of virus infection processes. By further comparing with the most widely used labeling strategy based on the Biotin-SA system, this new strategy has advantages of both high labeling efficiency and good retention of virus infectivity, thus offering a promising alternative for virus-labeling. Moreover, due to the ubiquitous presence of exposed amino groups on the surface of various viruses, this selective, efficient, reproducible and biofriendly strategy should have good universality for labeling both enveloped and nonenveloped viruses.

The size of C-nanodots can be electrochemically tuned by changing the applied potential during their preparation. The higher the applied potential, the smaller the resulting C-nanodots. Moreover, the surface oxidation degree of the C-nanodots can also be electrochemically tuned. The red-shift of emission independent of the size provides an insight into the luminescence mechanism of C-nanodots.

The prognostic features of T1N0M0 renal cell carcinoma (RCC) in Asian patients have not been well explored in large sample studies. In this study, we retrospectively analyzed the records of 713 patients undergoing nephrectomy for T1N0M0 RCC between 1991 and 2009 in three Asian hospitals. Univariate and multivariate analysis were performed to identify the independent predictive factors for T1N0M0 RCC prognosis among a series of Clinicopathological parameters, including age, gender, tumor size, Fuhrman grade, and histological classification. Our results showed that 388 of 713 patients had tumors 4.0 cm or smaller (stage T1a) and 325 of 713 patients had tumors 4.0–7.0 cm in size (stage T1b). Five-year cancer-specific survival (CSS) and recurrence-free survival (RFS) rates for this group of patients were 96.0% and 93.5%, respectively. The patients with T1b RCC had a significantly lower 5-year CSS and RFS rates than did those with T1a RCC (CSS, 93.1% vs. 98.6%, P = 0.026; RFS, 90.0% vs. 96.5%, P < 0.001). Patients with low grade (grades I–II) tumors had a higher 5-year CSS (97.8% vs. 91.2%, P = 0.001) and RFS (95.5% vs. 85.5%, P < 0.001) rate than did those with high grade (grades I–II) tumors. More interestingly, when stratifying patients to T1a and T1b groups, the role of grade in distinguishing prognosis could be only observed in patients with T1b disease. Cox regression showed tumor size and Fuhrman grade were significant in predicting CSS and RFS. Our study suggests that the prognosis of patients with T1N0M0 RCC is excellent, and these results are comparable to previously reported studies in Western patients. Furthermore, our data indicates that patients with T1b disease and high Fuhrman grade have high risk of tumor recurrence and death, thus requiring more frequent follow-up.

A strategy for construction of a solid-state electrogenerated chemiluminescence (ECL) sensor based on a self-assembled film of core/shell structural CdSe/ZnS quantum dots (QDs) is presented in this work for the first time. The surface coverage of CdSe/ZnS QDs is about 2.2 × 1012 NPs/cm2, which is on the level of a submonolayer. This self-assembled CdSe/ZnS QD film has shown a strong and efficient cathodic ECL with peroxydisulfate and hydroperoxide as coreactants in aqueous solution. The quenching effect with hydroperoxide as the coreactant reveals an energy-level-related mechanism for the QD ECL processes. The energy levels of QDs and coreactants determine ECL responses. This insight provides a universal guide to select proper coreactants for the QD ECL sensors. In addition, the self-assembled CdSe/ZnS QD ECL sensor has the potential to detect energy-level-related analytes, such as dopamine.

This study was carried out to assess whether the spatial resolution has an impact on the detection accuracy of proximal caries in flat panel CBCT (cone beam computerized tomography) images and if the detection accuracy can be improved by flat panel CBCT images scanned with high spatial resolution when compared to digital intraoral images. The CBCT test images of 45 non-restored human permanent teeth were respectively scanned with the ProMax 3D and the DCT Pro scanners at different resolutions. Digital images were obtained with a phosphor plate imaging system Digora Optime. Eight observers evaluated all the test images for carious lesion within the 90 proximal surfaces. With the histological examination serving as the reference standard, observer performances were evaluated by receiver operating characteristic (ROC) curves. The areas under the ROC curves were analyzed with two-way analysis of variance. No significant differences were found among the CBCT images and between CBCT and digital images when only proximal enamel caries was detected (p = 0.989). With respect to the detection of proximal dentinal caries, significant difference was found between CBCT and digital images (p < 0.001) but not among CBCT images. The spatial resolution did not have an impact on the detection accuracy of proximal caries in flat panel CBCT images. The flat panel CBCT images scanned with high spatial resolution did not improve the detection accuracy of proximal enamel caries compared to digital intraoral images. CBCT images scanned with high spatial resolutions could not be used for proximal caries detection.

This study presents the investigation of bioconjugating ability of near-infrared (NIR) CdSeTe/ZnS quantum dots (QDs) (710 nm) and visible CdSe QDs (595 nm) in immunofluorescent staining for cancer biomarkers in gastric cancer tissues probed with the homemade Hadamard transform (HT) spectral imaging microscope and a commercial multispectral imaging system. The results show that imunostaining ability of NIR QDs probes is stronger than that of visible QDs when the two kinds of QDs are simultaneously used to probe the cancer biomarkers such as cytokeratin 20 (CK20) and proliferating cell nuclear antigen (PCNA) in gastric cancer tissues. Moreover, when the two QDs probes are used for immunostaining successively for the same target molecules, staining order has great influences on the final results due to their different conjugating ability to the marker proteins. The results imply that NIR QDs hold more promise for real-time imaging of tumor tissues due to its higher sensitivity and contrast. In addition, the results also demonstrate the potential of Hadamard transform spectral imaging as a useful tool in biomedical analysis and quantitative evaluation for tumor tissues.

Introduction The objective of this study was to assess the clinical outcome after removal of a broken file from root canal by using ultrasonic techniques and to evaluate various factors that might influence the outcome. Methods One hundred two patients with 102 teeth with a broken file in the root canal, treated between 2004 and 2008, were contacted for follow-up examination 12–68 months after treatment (median, 28 months). Outcome was assessed on the basis of clinical and radiographic criteria by calibrated examiners and dichotomized as healed or disease. The result was analyzed statistically by Pearson or Fisher exact test and multivariate logistic regression. Results Of 66 teeth examined (64.7% recall), 81.8% were healed. Stepwise logistic regression analysis revealed that inadequate root canal filling was a statistically significant factor in predicting failure of successful healing. The cases with removal of broken files had a higher healing rate than those with broken files left in the root canal, but the difference was not statistically significant ( P > .05). There was no significant difference in outcome related to presence or absence of perforation, although the rate of healing was greater in teeth without perforation. Other factors including age, sex, tooth type, apical status, position of segment, and type of restoration did not affect prognosis. Conclusions Treatment outcome after the use of ultrasonics for removal of broken files was favorable. Adequate root filling has a more substantial impact on treatment outcome than the other factors.

Accurate and rapid recognition and separation of multiple types of biological targets such as molecules, cells, bacteria or viruses from complex sample mixtures is of great importance for a wide range of diagnostic and therapeutic strategies. To achieve this goal, a set of fluorescent, magnetic, dual-encoded multifunctional bioprobes has been constructed by co-embedding different-sized quantum dots and varying amounts of γ-Fe(2)O(3) magnetic nanoparticles into swollen poly(styrene/acrylamide) copolymer nanospheres. The dual-encoded bioprobes, which possessed different photoluminescent property and magnetic susceptibility, were proven to be capable of simultaneously recognizing and separating multiple components from a complex sample when three kinds of lectins were used as the targets. The lectins were separated with high efficiency and kept their bioactivity during the process. Compared to the conventional batchwise separation, this method does not require a large number of sequential reaction steps, which is economical of time and can be very reagent-saving. By combining the multiplexing capability of quantum dots with the superparamagnetic properties of iron oxide nanoparticles, this dual-encoded technique is expected to open new opportunities in high-throughput and multiplex bioassays, such as cell sorting, proteomical and genomical applications, drug screening etc.

Fluorescent-magnetic-biotargeting multifunctional nanobioprobes (FMBMNs) have attracted great attention in recent years due to their increasing, important applications in biomedical research, clinical diagnosis, and biomedicine. We have previously developed such nanobioprobes for the detection and isolation of a single kind of tumor cells. Detection and isolation of multiple tumor markers or tumor cells from complex samples sensitively and with high efficiency is critical for the early diagnosis of tumors, especially malignant tumors or cancers, which will improve clinical diagnosis outcomes and help to select effective treatment approaches. Here, we expanded the application of the monoclonal antibody (mAb)-coupled FMBMNs for multiplexed assays. Multiple types of cancer cells, such as leukemia cells and prostate cancer cells, were detected and collected from mixed samples within 25 minutes by using a magnet and an ordinary fluorescence microscope. The capture efficiencies of mAb-coupled FMBMNs for the above mentioned two types of cells were 96% and 97% respectively. Furthermore, by using the mAb-coupled FMBMNs, specific and sensitive detection and rapid separation of a small number of spiked leukemia cells and prostate cancer cells in a large population of cultured normal cells (about 0.01% were tumor cells) were achieved simply and inexpensively without any sample pretreatment before cell analysis. Therefore, mAb-coupled multicolour FMBMNs may be used for very sensitive detection and rapid isolation of multiple cancer cells in biomedical research and medical diagnostics.

Objectives To evaluate the accuracy of cone-beam computerized tomography (CBCT) for the detection of noncavitated proximal caries and to compare the detection accuracies of 2 CBCT imaging systems with those based on plain-film radiographs and phosphor-plate images. Study design Test radiographs of 39 noncavitated unrestored human permanent teeth were obtained with film, phosphor-plate, ProMax 3D, and Kodak 9000 3D imaging systems. Seven observers used a 5-level scale to evaluate test images for the presence of proximal caries. With histologic examination serving as the reference standard, observer performances were assessed with receiver operating characteristic (ROC) curves, and the areas under the ROC curves (Az values) for the observers, and modalities were analyzed with a repeated-measures analysis of variance. Results The mean Az values for film, phosphor plates, ProMax 3D, and Kodak 9000 3D imaging systems were 0.541, 0.523, 0.528, and 0.525, respectively (P = .763). Conclusion For detecting subtle noncavitated proximal caries, the detection accuracy with the CBCT images was little better than chance performance and was similar to that with phosphor plate– and film-based intraoral images.

The aim of the present study was to assess the treatment outcomes in a cohort of adult patients with intratesticular rhabdomyosarcoma. Between 1999 and 2008, 296 patients underwent radical orchiectomy for intrascrotal mass. A retrospective chart review was carried out for adult patients diagnosed with intratesticular rhabdomyosarcoma. Overall, six patients (mean age 21 years, range 17-23) were included: five had embryonic rhabdomyosarcoma and one had pleomorphic rhabdomyosarcoma. Four patients underwent retroperitoneal lymph node dissection and five patients had postoperative chemotherapy. The mean length of follow up was 28 months (range 12-51 months). One patient was lost to follow up after 26 months, five cases remained alive at the end of the study. Surgical resection with chemotherapy was an effective way to achieve favorable outcomes and long-term survival in adult patients with metastases from intratesticular rhabdomyosarcoma.

Quantum dots (QDs) are inorganic fluorescent nanocrystals with excellent properties such as tunable emission spectra and photo-bleaching resistance compared with organic dyes, which make them appropriate for applications in molecular beacons. In this work, quantum dot-based molecular beacons (QD-based MBs) were fabricated to specifically detect β-lactamase genes located in pUC18 which were responsible for antibiotic resistance in bacteria Escherichia coli (E. coli) DH5α. QD-based MBs were constructed by conjugating mercaptoacetic acid-quantum dots (MAA-QDs) with black hole quencher 2 (BHQ2) labeled thiol DNA vial metal-thiol bonds. Two types of molecular beacons, double-strands beacons and hairpin beacons, were observed in product characterization by gel electrophoresis. Using QD-based MBs, one-step FISH in tiny bacteria DH5α was realized for the first time. QD-based MBs retained their bioactivity when hybridizing with complementary target DNA, which showed excellent advantages of eliminating background noise caused by adsorption of non-specific bioprobes and achieving clearer focus of genes in plasmids pUC18, and capability of bacterial cell penetration and signal specificity in one-step in situ hybridization.

By kinetically controlling a biomimetic reduction in a quasi-biological system (an aqueous solution containing electrolyte, peptide, and coenzyme), watersoluble, glutathione-capped Au clusters with a mean diameter of 1.3 nm are successfully synthesized via a new route. Opportunities that facilitate the control of the reaction are created in such a quasi-biological system. The relatively slow rate of the biomimetic reduction, the pH-dependent reducibility of the reducing agent, and the favorable structure of the capping molecules conspire together for the realization of a kinetics-controlled formation of the Au clusters under mild conditions. Compared to existing methods of synthesizing gold clusters by stoichiometrically controlling the molar ratio of the Au atoms and the ligands, our current method is based on slowing down the reduction of the Au precursors in the initial stage by adjusting the activity of the reducing agent in real time, instead of using a strong reducing agent such as NaBH 4 . This strategy of rationally utilizing biological or biomimetic processes with unique features to provide a benefi cial complement to conventional chemical syntheses could open a new way for the sustainable development of nanotechnology.

A simple, controllable and effective sample preparation method was established for atomic force microscopy (AFM) imaging of individual DNA molecules in aqueous solution. Firstly, magnesium ion (Mg(2+)) at a concentration of 5.0-10.0 mM as a positively charged bridge was transferred onto mica to immobilize DNA molecules. Then Mg(2+)-modified mica was used to investigate DNA molecules in any buffer without magnesium ion by AFM. AFM images demonstrated that DNA molecules can be successfully observed in solution with good resolution, reproducibility, and stability. Further, this DNA sample preparation method makes AFM successful to investigate DNA molecular interaction in situ and DNA/chitosan complex in gene delivery.

Objective: To analyze the incidence of benign lesions in Chinese patients undergoing nephrectomies for renal masses identified as localized renal cell carcinoma (RCC) in preoperative imaging. Methods: Between 1999 and 2007, 303 patients (112 female, 191 male) with presumed localized RCC underwent nephrectomy (234 radical nephrectomies and 69 partial nephrectomies). Preoperative computed tomography images and pathological findings were reviewed and analyzed. Results: Pathological examinations revealed 31 (10.2%) benign lesions in the 303 patients. Among these 31 benign lesions, 15 (5.0%) were angiomyolipomas (AML) and only four (1.3%) were oncocytomas. Significantly, 20 (17.9%) of the 112 female patients had benign lesions compared with 11 (5.8%; P = 0.001) male patients. Benign renal lesions were found in five (25.0%) of the 20 patients with renal masses smaller than 2 cm, 13 (13.0%) of the 100 patients with renal masses 2–4 cm in size and 13 (7.1%) of the 183 patients with renal masses larger than 4 cm. Conclusions: Patients in the present study population show a low incidence of benign renal lesions, approximately half of them being AML. Female patients and patients with renal masses smaller than 4 cm are more likely to have benign renal lesions.

Ag2 S quantum dots (QDs) are well-known near-infrared fluorophores and have attracted great interest in biomedical labeling and imaging in the past years. However, their photoluminescence efficiency is hard to compete with Cd-, Pb-based QDs. The high Ag+ mobility in Ag2 S crystal, which causes plenty of cation deficiency and crystal defects, may be responsible mainly for the low photoluminescence quantum yield (PLQY) of Ag2 S QDs. Herein, a cation-doping strategy is presented via introducing a certain dosage of transition metal Pb2+ ions into Ag2 S nanocrystals to mitigate this intrinsic shortcoming. The Pb-doped Ag2 S QDs (designated as Pb:Ag2 S QDs) present a renovated crystal structure and significantly enhanced optical performance. Moreover, by simply adjusting the levels of Pb doping in the doped nanocrystals, Pb:Ag2 S QDs with bright emission (PLQY up to 30.2%) from 975 to 1242 nm can be prepared without altering the ultrasmall particle size (≈2.7-2.8 nm). Evidently, this cation-doping strategy facilitates both the renovation of crystal structure of Ag2 S QDs and modulation of their optical properties.

Hydrothermal reactions of 1,2,4-triazole (Htrz) with CuBr or CuCl2 in the presence of benzene-1,3,5-tricarboxylic acid (H3btc) produced the infinite aggregates, [CuI4CuII2(trz)4(btc)Br] (1) and [CuI4CuII2(trz)4(btc)Cl] (2), composed of 2D mixed valent Cu-trz sheets, benzene-1,3,5-tricarboxylates and halides; in particular, the Br-labelled 1 crystallizes in I4/m space group with cell parameters: a = 20.321(2) A, b = 20.321(2) A, c = 13.699(1) A, V = 5714.8(4) A3 and the Cl-labelled 2 in a P4/ncc with a = 20.3711(5) A, b = 20.3711(5) A, c = 13.7711(8) A, V = 5656.9(6) A3. Solid measurements reveal that the halide-discriminated compounds are thermostable up to 300 °C and ambient luminescent, exhibiting the emission bands in the range of 400–550 nm.

To assess the impact of surgical approaches and clinico-pathological parameters on the prognosis of localized renal cell carcinoma (RCC) after laparoscopic radical nephrectomy (LRN) or open radical nephrectomy (ORN). Between 2002 and 2008, 336 patients with localized RCC were treated at our two institutions. Of these, 261 were classified as clinical stage pT1 and 75 as pT2, the mean age was 52.3 years. One hundred forty-two patients underwent LRN, and 194 underwent ORN. The medical records of all patients were retrospectively reviewed. The mean follow-up was 44 months (12–84 months). The 3-, 5-, and 7-year cancer-specific survival (CSS) rates of LRN group (96.3, 94.6, and 92.5%, respectively) were equivalent to ORN (97.4, 94.2, and 91.4%, respectively). The mean CSS rates were not significantly different between LRN and ORN groups for either pT1 (82.3 vs. 81.6 months, P = 0.713) or pT2 (69.0 vs. 72.1 months, P = 0.729). Univariate analysis demonstrated significant impact of tumor stage (pT1 vs. pT2, P = 0.002) and tumor grade (G1/2 vs. G3/4, P = 0.045) on CSS. Surgical approach (laparoscopic or open) was not a statistically significant factor on CSS. Multivariate analysis identified that tumor stage was an independent prognostic factor for CSS of localized RCC (P = 0.007). Based on the long-term follow-up, our evaluation suggests that LRN is oncologically equivalent to ORN for patients with pT1 or pT2 tumors. Among the clinical variables, tumor stage appears to be an independent prognostic factor of CSS of localized RCC patients.

To introduce a modified technique for urethral anastomosis in orthotopic neobladder reconstruction.Between January 2002 and August 2008, about 141 consecutive patients (130 men and 11 women) underwent total cystectomy and orthotopic neobladder reconstruction in which a modified technique was used to anastomosed the caudal-most part of the intestinal neobladder directly to the urethral remnant. The emptying of the neobladder, the early and late complications at the urethral anastomosis was evaluated.Mean patient age at surgery was 58.4 years (range, 33-83) and median follow-up was 42 months (range, 4-83 months). Early urine leakage at the vesicourethral anastomosis developed in 1 patient, and was cured by extending catheter drainage. A total of 138 patients had good emptying of the neobladder, with residual urine volume50 mL. Three patients had residual urine volume100 mL, and achieved good emptying after intermittent catheterization once a week for 6-12 months. No late complications occurred at the urethral anastomosis site. Daytime continence was good or satisfactory in 97.0% of patients and night-time continence was good or satisfactory in 88.5% of patients.The clinical outcome of our modified technique for urethral anastomosis in orthotopic neobladder substitution was excellent, but the advantage of this technique needs prospective controlled study.

Spermatogonial stem cells (SSCs) divide continuously to support spermatogenesis throughout postnatal life and transmit genetic information to the next generation. Here, we report the successful establishment of the method for the isolation and identification of human SSCs from testicular tissue, and to determine the culture conditions required to expand SSCs on human embryonic stem cell-derived fibroblast-like cells (hdFs). Large-scale cultures of SSCs were maintained on hdF feeder layers and expanded in the presence of a combination of cytokines and glial cell line-derived neurotrophic factor for at least 2 months. Cell surface marker analysis showed that SSCs retained high levels of alkaline phosphatase activity and stained strongly for anti-stage-specific embryonic antigen (SSEA)-1, OCT4 and CD49f. They also expressed the genes OCT4, SOX3 and STRA8 as detected by reverse transcription polymerase chain reaction (RT-PCR) analysis. These data clearly illustrate a novel approach for the growth of human SSCs using hdFs as feeder cells, potentially eliminating xenogeneic contaminants. This system provides a new opportunity for the study of the regulatory mechanism of the 'niche' that governs SSC self-renewal, and will be a valuable source of SSCs for potential clinical applications.

There are currently some problems in the field of chemical synthesis, such as environmental impact, energy loss, and safety, that need to be tackled urgently. An interdisciplinary approach, based on different backgrounds, may succeed in solving these problems. Organisms can be chosen as potential platforms for materials fabrication, since biosystems are natural and highly efficient. Here, an example of how to solve some of these chemical problems through biology, namely, through a novel biological strategy of coupling intracellular irrelated biochemical reactions for controllable synthesis of multicolor CdSe quantum dots (QDs) using living yeast cells as a biosynthesizer, is demonstrated. The unique fluorescence properties of CdSe QDs can be utilized to directly and visually judge the biosynthesis phase to fully demonstrate this strategy. By such a method, CdSe QDs, emitting at a variety of single fluorescence wavelengths, can be intracellularly, controllably synthesized at just 30°C instead of at 300°C with combustible, explosive, and toxic organic reagents. This green biosynthetic route is a novel strategy of coupling, with biochemical reactions taking place irrelatedly, both in time and space. It involves a remarkable decrease in reaction temperature, from around 300 °C to 30 °C and excellent color controllability of CdSe photoluminescence. It is well known that to control the size of nanocrystals is a mojor challenge in the biosynthesis of high-quality nanomaterials. The present work demonstrates clearly that biological systems can be creatively utilized to realize controllable unnatural biosynthesis that normally does not exist, offering new insights for sustainable chemistry.

Ferrichrome is a kind of high-affinity iron(III) chelators and its complex with Fe3+ can be specifically recognized by the corresponding receptors located on the outer membrane of some bacterial ce...

Quantum dots (QDs) have been used as a new class of bioprobes in medical imaging in recent years. The study of interaction between QDs and biomacromolecules is important for interpreting biological data. In this work, Rayleigh light scattering (RLS) was employed to investigate the nonspecific interaction between mercaptoacetic acid modified CdSe/ZnS quantum dots (MAA-QDs) and human immunoglobulin G (IgG). The conjugation processes between QDs and IgG in different conditions including addition sequence, pH were carefully studied. The addition of IgG to QDs solution was found to form a fixed size of QDs-IgG conjugate, with the QDs-to-IgG ratio of approximately 13, while the addition of QDs to IgG solution resulted in a gradually increased conjugate size, with variable QDs-to-IgG ratio till the binding saturation was reached.

We report on a simple method for self loading and culture of mammalian cells in microfluidic multi-chambers for high throughput screening. The device was obtained by using one layer soft lithography with polydimethylsiloxane (PDMS) and thermal bonding on a glass slide. Self loading of cell suspension could be possible after degassing of the PDMS device for 30 min. Both cell loading efficiency and cell proliferation behaviors have been analyzed with triangle chambers of different sizes, all connected to the main flow channels with small entrances. We found that the number of cells loaded into the micro-chamber increased with the side length of the triangle, showing well size dependence and that self loading at a single cell level was possible for small chambers. For large chambers, the cell area density after loading and proliferation is however quite heterogeneous. For demonstration, HeLa cell growth behavior has been followed for 11 days until the total area of the largest chambers was fully filled.