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Background Magnetic nanoparticles (MNPs) can be localized against hemodynamic forces in blood vessels with the application of an external magnetic field. In addition, PEGylation of nanoparticles may increase the half-life of nanocomposites in circulation. In this work, we examined the effect of PEGylation on the magnetic capture of MNPs in vivo. Methods Laser speckle contrast imaging and capillaroscopy were used to assess the magnetic capture of dextran-coated MNPs and red blood cell (RBC) flow in cremaster microvessels of anesthetized rats. Magnetic capture of MNPs in serum flow was visualized with an in vitro circulating system. The effect of PEGylation on MNP-endothelial cell interaction was studied in cultured cells using an iron assay. Results In microcirculation through cremaster muscle, magnet-induced retention of 250 nm MNPs was associated with a variable reduction in RBC flow, suggesting a dynamic coupling of hemodynamic and magnetic forces. After magnet removal, faster restoration of flow was observed in PEG(+) than PEG(-) group, which may be attributed to a reduced interaction with vascular endothelium. However, PEGylation appears to be required for magnetic capture of 50 nm MNPs in microvessels, which was associated with increased hydrodynamic diameter to 130±6 nm in serum, but independent of the ς-potential. Conclusion These results suggest that PEGylation may enhance magnetic capture of smaller MNPs and dispersion of larger MNPs after magnet removal, which may potentially affect the targeting, pharmacokinetics and therapeutic efficacy.

Dengue virus (DENV) circulates in tropical and subtropical areas around the world, where it causes high morbidity and mortality. There is no effective treatment of infection, with supportive care being the only option. Furthermore, early detection and diagnosis are important to facilitate clinical decisions. In this study, seven monoclonal antibodies (mAbs) recognizing nonstructural protein 1 (NS1) of DENV were generated by hybridoma techniques. These antibodies can be divided into two groups: serotype-specific (DB6-1, DB12-3, and DB38-1) and nonspecific (consisting of antibodies DB16-1, DB20-6, DB29-1, and DB41-2). The B-cell epitopes of DB20-6 and DB29-1 were identified by phage display and site-directed mutagenesis, and its binding motif, WXXWGK, was revealed to correspond to amino acid residues 115-120 of the DENV-2 NS1 protein. A diagnostic platform, consisting of a serotype-specific capture antibody and a complex detection antibody, exhibited a detection limit of about 1 ng/mL, which is sufficient to detect NS1 in clinical serum samples from dengue patients. This diagnostic platform displayed better specificity and sensitivity than two examined commercial NS1 diagnostic platforms. In summary, our results indicate that these newly generated mAbs are suitable for detection of NS1 protein of DENV-2 in clinical samples.

Background Magnetic nanoparticles (MNPs) can be localized against hemodynamic forces in blood vessels with the application of an external magnetic field. In addition, PEGylation of nanoparticles may increase the half-life of nanocomposites in circulation. In this work, we examined the effect of PEGylation on the magnetic capture of MNPs in vivo. Methods Laser speckle contrast imaging and capillaroscopy were used to assess the magnetic capture of dextran-coated MNPs and red blood cell (RBC) flow in cremaster microvessels of anesthetized rats. Magnetic capture of MNPs in serum flow was visualized with an in vitro circulating system. The effect of PEGylation on MNP-endothelial cell interaction was studied in cultured cells using an iron assay. Results In microcirculation through cremaster muscle, magnet-induced retention of 250 nm MNPs was associated with a variable reduction in RBC flow, suggesting a dynamic coupling of hemodynamic and magnetic forces. After magnet removal, faster restoration of flow was observed in PEG(+) than PEG(–) group, which may be attributed to a reduced interaction with vascular endothelium. However, PEGylation appears to be required for magnetic capture of 50 nm MNPs in microvessels, which was associated with increased hydrodynamic diameter to 130±6 nm in serum, but independent of the ς-potential. Conclusion These results suggest that PEGylation may enhance magnetic capture of smaller MNPs and dispersion of larger MNPs after magnet removal, which may potentially affect the targeting, pharmacokinetics and therapeutic efficacy.

Dengue virus (DENV) infection is currently at pandemic levels, with populations in tropical and subtropical regions at greatest risk of infection. Early diagnosis and management remain the cornerstone for good clinical outcomes, thus efficient and accurate diagnostic technology in the early stage of the disease is urgently needed. Serotype-specific monoclonal antibodies (mAbs) against the DENV1 nonstructural protein 1 (NS1), DA12-4, DA13-2, and DA15-3, which were recently generated using the hybridoma technique, are suitable for use in diagnostic platforms. Immunofluorescence assay (IFA), enzyme-linked immunosorbent assay (ELISA) and Western blot analysis further confirmed the serotype specificity of these three monoclonal antibodies. The ELISA-based diagnostic platform was established using the combination of two highly sensitive mAbs (DA15-3 and DB20-6). The same combination was also used for the flow cytometry-based diagnostic platform. We report here the detection limits of flow cytometry-based and ELISA-based diagnostic platforms using these mAbs to be 0.1 and 1 ng/mL, respectively. The collected clinical patient serum samples were also assayed by these two serotyping diagnostic platforms. The sensitivity and specificity for detecting NS1 protein of DENV1 are 90% and 96%, respectively. The accuracy of our platform for testing clinical samples is more advanced than that of the two commercial NS1 diagnostic platforms. In conclusion, our platforms are suitable for the early detection of NS1 protein in DENV1 infected patients.

A chip-level optical interconnect module combined with a vertical-cavity surface-emitting laser (VCSEL) chip, a photodetector (PD) chip, a driver integrated circuit (IC), and an amplifier IC on a silicon-on-insulator (SOI) substrate with 3-D guided-wave paths is experimentally demonstrated. Such an optical interconnect is developed for the signal connection in multicore processors or memory-to-processor interfaces. The 3-D guided-wave path, consisting of silicon-based 45 $^{\circ}$ microreflectors and trapezoidal waveguides, is used to connect the optical signal between transmitter and receiver. In this paper, the VCSEL and PIN PD chips are flip-chip integrated on a SOI substrate to achieve complete chip-level optical interconnects. Due to the unique 3-D guided-wave path design, a higher laser-to-PD optical coupling efficiency of $-$ 2.19 dB and a larger alignment tolerance of $\pm 10\ \mu\hbox{m}$ for the VCSEL/PD assembly are achieved. The measured laser-to-PD optical transmission efficiency can reach $-$ 2.19 dB, and the maximum optical power and threshold current of VCSEL is 3.27 mW and 1 mA, respectively. To verify the data transmission, the commercial driver IC and amplifier IC are assembled upon the silicon chip, and the error-free data transmission of 10 Gbps can be achieved when the VCSEL is operated at the driving current of 9 mA.

When people close eyes to relax, an alpha wave in the frequency range of 8–13 Hz appears from brain signals. There were many medical reports proofed that some specific music can resonate with the alpha wave and strengthen the wave. Therefore, this alpha wave music can improve more relaxing for people and are very helpful when they need to take a rest. Due to the alpha wave music is classified manually by experts only, it is not popular in the market currently. In this paper, we will investigate the content-based features of the alpha wave music and use them to analyze the similarity between alpha wave music and existing music genres. The purpose of this research is to find the music which is similar to alpha wave music, such that we can recommend to users for relaxing before the automatic classification scheme for alpha wave music being developed.

The chip-level 1 × 2 optical interconnects using the polymer vertical splitter developed on a silicon substrate are demonstrated. The 1 × 2 vertical-splitting configuration is realized using a polymer waveguide terminated at three silicon 45 ° reflectors. The high-frequency transmission lines combined with the indium solder bumps are developed to flip-chip assemble a vertical-cavity surface-emitting laser chip at the input port and two photodetector chips at two output ports. Total transmission loss of -3.26 dB with a splitting ratio of 1 : 1 for the proposed splitter is experimentally obtained. A 10-Gbit/s data transmission with bit error rates better than 10-12 for two output ports is achieved. It reveals that such chip-level 1 × 2 optical interconnects using the polymer vertical splitter are suitable for high-speed data transmission with multiple output ports.

It is known that solid tumors recruit new blood vessels to support tumor growth, but the molecular diversity of receptors in tumor angiogenic vessels might also be used clinically to develop better targeted therapy. In vivo phage display was used to identify peptides that specifically target tumor blood vessels. Several novel peptides were identified as being able to recognize tumor vasculature but not normal blood vessels in severe combined immunodeficiency (SCID) mice bearing human tumors. These tumor-homing peptides also bound to blood vessels in surgical specimens of various human cancers. The peptide-linked liposomes containing fluorescent substance were capable of translocating across the plasma membrane through endocytosis. With the conjugation of peptides and liposomal doxorubicin, the targeted drug delivery systems enhanced the therapeutic efficacy of the chemotherapeutic agent against human cancer xenografts by decreasing tumor angiogenesis and increasing cancer cell apoptosis. Furthermore, the peptide-mediated targeting liposomes improved the pharmacokinetics and pharmacodynamics of the drug they delivered compared with nontargeting liposomes or free drugs. Our results indicate that the tumor-homing peptides can be used specifically target tumor vasculature and have the potential to improve the systemic treatment of patients with solid tumors.

Severe acute respiratory syndrome (SARS) has emerged as a highly contagious, sometimes fatal disease. To find disease-specific B cell epitopes, phage-displayed random peptide libraries were panned on serum immunoglobulin (Ig) G antibodies from patients with SARS. Forty-nine immunopositive phage clones that bound specifically to serum from patients with SARS were selected. These phageborne peptides had 4 consensus motifs, of which 2 corresponded to amino acid sequences reported for SARS-associated coronavirus (SARSCoV). Synthetic peptide binding and competitive-inhibition assays further confirmed that patients with SARS generated antibodies against SARS-CoV. Immunopositive phage clones and epitope-based peptide antigens demonstrated clinical diagnostic potential by reacting with serum from patients with SARS. Antibody-response kinetics were evaluated in 4 patients with SARS, and production of IgM, IgG, and IgA were documented as part of the immune response. In conclusion, B cell epitopes of SARS corresponded to novel coronavirus. Our epitope-based serologic test may be useful in laboratory detection of the virus and in further study of the pathogenesis of SARS.

In this study, a serotype-specific monoclonal antibody (mAb), D216-1 (Ab4), against dengue virus type 2 (DEN-2) was generated. The specificity of Ab4, which recognized DEN-2 non-structural protein 1, was determined by ELISA, immunofluorescence and immunoblotting analyses. The serotype-specific B-cell epitope of Ab4 was identified further from a random phage-displayed peptide library; selected phage clones reacted specifically with Ab4 and did not react with other mAbs. Immunopositive phage clones displayed a consensus motif, His–Arg/Lys–Leu/Ile, and a synthetic peptide corresponding to the phage-displayed peptide bound specifically to Ab4. The His and Arg residues in this epitope were found to be crucial for peptide binding to Ab4 and binding activity decreased dramatically when these residues were changed to Leu. The epitope-based synthetic peptide not only identified serum samples from DEN-2-immunized mice and rabbits by ELISA but also differentiated clearly between serum samples from DEN-2- and Japanese encephalitis virus-immunized mice. This mAb and its epitope-based peptide antigen will be useful for serologic diagnosis of DEN-2 infection. Furthermore, DEN-2 epitope identification makes it feasible to dissect antibody responses to DEN and to address the role of antibodies in the pathogenesis of primary and secondary DEN-2 infections.

A whole on-chip optical interconnects integrated with laser, photodetectors, driver IC, and amplifier IC is experimentally demonstrated. A 10-Gbps error-free data transmission is achieved as driving current of laser is 10 mA.

The chip-level 10-Gbit/s optical interconnects with the BER better than 10-12 using the 1 × 2 polymer vertical splitter, which is composed of a polymer waveguide and three silicon 45° reflectors is demonstrated.

The heterotrimeric GTP-binding regulatory proteins (G proteins) play an important role in the regulation of membrane signal transduction. Recently, we identified the association of Go protein with mitotic spindles. Here we have investigated the relationship between Go protein and microtubules. We used temperature-dependent reversible assembly and taxol methods to purify microtubules from bovine brains. Goalpha and Gbeta proteins were identified in the microtubular fraction by both methods. The Goalpha subunit in the microtubular fraction could be ADP ribosylated by pertussis toxin. Co-immunoprecipitation data also revealed that Go protein can interact with microtubules. Exogenous Go protein could be incorporated into the assembled microtubular fraction, and 5 microg/ml (60 nM) of Go protein inhibited 40% of microtubule assembly. Western blot analysis of Goalpha-1 and Goalpha-2 in microtubular fractions showed that only Goalpha-1 is associated with microtubules. We conclude that the Goalpha-1betagamma proteins are associated with microtubules and may play some role in regulating the assembly and disassembly of microtubules.

The association of G protein β2 subunit (Gβ2) with mitotic spindles in various mammalian cells has been demonstrated previously. Recently, we have identified the association of Gβ2 protein with microtubules (Wu et al., [1998] J. Cell. Biochem. 70: 552-562). In the present experiment we have demonstrated the possible functional role of Gβ2 in microtubule and mitotic spindle organization in mammalian cells. When Gβ2 antisense phosphorothioate oligonucleotides were transfected into mammalian cells, inhibition of cell proliferation with cell death after a 4-day treatment was observed. If the transfected cells were incubated for two days and their Gβ2 and microtubules were examined by Western blotting and immunofluorescence localization, marked reduction of the Gβ2 protein, fragmentation and disassembly of cytoplasmic microtubules, and disorganized mitotic spindles were found. We conclude that the Gβ2 protein is closely associated with microtubule assembly and may play a potential role in the regulation of cell proliferation and microtubule and mitotic spindle organization in mammalian cells. J. Cell. Biochem. 83: 136–146, 2001. © 2001 Wiley-Liss, Inc.

Targeted delivery of drugs to tumors represents a significant advance in cancer diagnosis and therapy. Therefore, development of novel tumor-specific ligands or pharmaceutical nanocarriers is highly desirable. In this study, we utilized phage display to identify a new targeting peptide, SP90, which specifically binds to breast cancer cells, and recognizes tumor tissues from breast cancer patients. We used confocal and electron microscopy to reveal that conjugation of SP90 with liposomes enables efficient delivery of drugs into cancer cells through endocytosis. Furthermore, in vivo fluorescent imaging demonstrated that SP90-conjugated quantum dots possess tumor-targeting properties. In tumor xenograft and orthotopic models, SP90-conjugated liposomal doxorubicin was found to improve the therapeutic index of the chemotherapeutic drug by selectively increasing its accumulation in tumors. We conclude that the targeting peptide SP90 has significant potential in improving the clinical benefits of chemotherapy in the treatment and the diagnosis of breast cancer.

The identification of molecules that are down-regulated in malignant phenotype is important for understanding tumor biology and their role in tumor suppression. We compared the expression profile of four normal nasal mucosal (NNM) epithelia and a series of nasopharyngeal cancinoma (NPC) cell lines using cDNA microarray and confirmed the actual expression of the selected genes, and found osteoprotegerin (OPG) to be ubiquitously deficient in NPC cells. We also found OPG to be down-regulated in various cancer cell lines, including oral, cervical, ovarian, lung, breast, pancreas, colon, renal, prostate cancer, and hepatoma. Administration of recombinant OPG (rOPG) brought about a reduction in cancer cell growth through apoptotic mechanism. We generated eleven monoclonal antibodies (MAbs) against OPG to study OPG's expression and biological functions in cancer cells. OPG was detected in the tumor stromal regions, but not in the cancer cell per se in surgical specimens of liver cancer. Quantitative reverse transcription-polymerase chain reaction (Q-RT-PCR) revealed that OPG was down-regulated in NPC tissues compared with normal nasal polyp (NNP) tissues. In addition, we showed OPG silencing to be associated with promoter methylation as well as histone modifications. In OPG-silenced cancer cell lines, the OPG gene promoter CpG dinucleotides were highly methylated. Compared to normal cells, silenced OPG gene in cancer cells were found to have reduced histone 3 lysine 4 tri-methylation (H3K4me3) and increased histone 3 lysine 27 tri-methylation (H3K27me3). Taken together, these results suggest that OPG silencing in carcinoma cancer cells occurs through epigenetic repression.

An increasing number of private and public organizations and educational institutions are incorporating Computer-Assisted Language Learning (CALL) technology into either their traditional classroom setting, or online English for Specific Purposes (ESP) training programs. In the role of facilitating students' learning, it is important for all stakeholders of ESP training programs to investigate the effectiveness of implementing online learning CALL systems into the distance learning environment and the traditional classroom environment of the programs. In order to determine the “effectiveness” of a corporate ESP training program, the approach for this study was to evaluate trainees' pretest and posttest scores related to the ESP training program. The experimental group of this one group pretest and posttest design study was a group of 18 Chinese adult male trainees enrolled in a flight academy's Aviation English training program that implemented with online learning CALL technology blended with an instructor, in central Florida. The intervention of the study was the implementation of online learning CALL technology blended with an instructor in the classroom environment. The length of the intervention was eight weeks of Aviation English training that implemented blended learning instructions. In addition, a survey instrument was developed to collect data on students' basic information, attitudes toward learning English with CALL technology, motivations for study English, and their perceptions of CALL technology as facilitating interactions among students. The surveys were completed by the students before and after two months of intervention. The study found that within two months of the implementation of the blended learning in the Aviation English training program, participants had significant improvement on their test scores. Participants in the study generally had positive attitudes toward learning English with CALL, before and after two months of the Aviation English training program. They also had positive perceptions of CALL technology in facilitating interactions in the classroom, before and after the training program. The study also found that participants who had positive perceptions of CALL in facilitating interactions in the classroom tended to have positive attitudes toward using CALL in learning English.

Background Dengue virus (DENV) is a significant public health threat in tropical and subtropical regions of the world. A therapeutic antibody against the viral envelope (E) protein represents a promising immunotherapy for disease control. Methodology/Principal Findings We generated seventeen novel mouse monoclonal antibodies (mAbs) with high reactivity against E protein of dengue virus type 2 (DENV-2). The mAbs were further dissected using recombinant E protein domain I-II (E-DI-II) and III (E-DIII) of DENV-2. Using plaque reduction neutralization test (PRNT) and mouse protection assay with lethal doses of DENV-2, we identified four serotype-specific mAbs that had high neutralizing activity against DENV-2 infection. Of the four, E-DIII targeting mAb DB32-6 was the strongest neutralizing mAb against diverse DENV-2 strains. Using phage display and virus-like particles (VLPs) we found that residue K310 in the E-DIII A-strand was key to mAb DB32-6 binding E-DIII. We successfully converted DB32-6 to a humanized version that retained potency for the neutralization of DENV-2 and did not enhance the viral infection. The DB32-6 showed therapeutic efficacy against mortality induced by different strains of DENV-2 in two mouse models even in post-exposure trials. Conclusions/Significance We used novel epitope mapping strategies, by combining phage display with VLPs, to identify the important A-strand epitopes with strong neutralizing activity. This study introduced potential therapeutic antibodies that might be capable of providing broad protection against diverse DENV-2 infections without enhancing activity in humans., Author Summary Dengue virus (DENV) infection remains a serious health threat despite the availability of supportive care in modern medicine. Monoclonal antibodies (mAbs) of DENV would be powerful research tools for antiviral development, diagnosis and pathological investigations. Here we described generation and characterization of seventeen mAbs with high reactivity for E protein of DENV. Four of these mAbs showed high neutralizing activity against DENV-2 infection in mice. The monoclonal antibody mAb DB32-6 showed the strongest neutralizing activity against diverse DENV-2 and protected DENV-2-infected mice against mortality in therapeutic models. We identified neutralizing epitopes of DENV located at residues K310 and E311 of viral envelope protein domain III (E-DIII) through the combination of biological and molecular strategies. Comparing the strong neutralizing activity of mAbs targeting A-strand with mAbs targeting lateral ridge, we found that epitopes located in A-strand induced stronger neutralizing activity than those located on the lateral ridge. DB32-6 humanized version was successfully developed. Humanized DB32-6 variant retained neutralizing activity and prevented DENV infection. Understanding the epitope-based antibody-mediated neutralization is crucial to controlling dengue infection. Additionally, this study also introduces a novel humanized mAb as a candidate for therapy of dengue patients.