Antibody affinity maturation using bacterial surface display

A drug-resistant cell line (EAC/Dox) was developed by repeated exposure of Ehrlich ascites carcinoma cells to Doxorubicin (Dox) in vivo in male albino Swiss mice (6-8 weeks old). The weekly i.p. injections of Dox to mice (2 or 4 mg/kg/week for 4 months) gave rise to Dox-resistant cell line EAC/Dox, which displayed typical multidrug resistant (MDR) features of cross-resistance to a number of structurally and functionally unrelated drugs like doxorubicin, vinblastine and cisplatin. Moreover, the EAC/Dox cell line had lower drug accumulation than drug-sensitive (EAC/S) cells. Study of Western blots and immunofluorescence revealed that P-glycoprotein 170 kDa (P-gp) was absent in EAC/Dox cells. The drug resistance appeared to be due to the presence of a higher level of reduced glutathione (GSH) and glutathione S-transferase (GST) in EAC/Dox cells than in drug-sensitive (EAC/S) cells. The two structurally similar hydroxamic acid derivatives, i.e. oxalyl bis(N-phenyl)hydroxamic acid (X1) and succinyl bis(N-phenyl)hydroxamic acid (X2), having very low in vitro toxicity (IC50 value 250 microg/ ml), were investigated for their efficacy to reverse MDR. The compound X1 was able to reverse the effect of MDR and reduce GST in EAC/Dox cells. The compound X2 had no ability to reverse the effect of MDR. Further study on the mechanism of glutathione depletion and the resistance modifying property of X1 on other cell lines is warranted.     

Characterization of microbial trophic structures of two anaerobic bioreactors processing sulfate-rich waste streams

A multi-compartment anaerobic bioreactor, designated the anaerobic migrating blanket reactor (AMBR), did not perform well in terms of chemical oxygen demand (COD) removal after an increase in sulfate load, compared to a conventional upflow anaerobic sludge blanket (UASB) reactor. The trophic structures of the bioreactors were characterized by analyzing the electron flows, formation and consumption of fermentation intermediates and terminal product (methane and hydrogen sulfide) formation. Critical performance parameters were linked to operational perturbations such as increase in sulfate load and changes in flow reversal schemes in the AMBR. Both of these manipulations affected the microbial communities, which were monitored by terminal restriction fragment length polymorphism (T-RFLP) analysis targeting the bacterial and archaeal domains. The less stable AMBR did not produce granular biomass, and in response to increased sulfate concentrations, experienced a reversal in the distribution of hydrogenotrophic methanogens that correlated with a shift in electron flow from butyrate to propionate. As this shift occurred, bacterial populations such as butyrate-producing clostridia, became predominant, thus leading to reactor imbalance. The stable UASB reactor developed and retained granules and maintained a relatively stable archaeal community. Sulfate perturbation led to the selection of a novel bacterial group (Thermotogaceae), which was most likely well adapted to the increasingly sulfidogenic conditions in the bioreactor.     

Evaluation of the corrosion resistance of anodized aluminum 6061 using electrochemical impedance spectroscopy (EIS)

The corrosion resistance of anodized Al 6061 produced by two different anodizing and sealing processes was evaluated using electrochemical impedance spectroscopy (EIS). The scanning electron microscope (SEM) was employed to determine the surface structure and the thickness of the anodized layers. The EIS data revealed that there was very little change of the properties of the anodized layers for samples that were hard anodized in a mixed acid solution and sealed in hot water over a 365 day exposure period in a 3.5 wt% NaCl solution. The specific admittance A_s and the breakpoint frequency f_b remained constant with exposure time confirming that the hard anodizing process used in this study was very effective in providing excellent corrosion resistance of anodized Al 6061 over extended exposure periods. Some minor degradation of the protective properties of the anodized layers was observed for samples that were hard anodized in H₂SO₄ and exposed to the NaCl solution for 14 days.     

A 10-Gb/s laser/modulator driver IC with a dual-mode activelymatched output buffer

A laser/modulator driver IC for 10-Gb/s-SONET OC-192-fiber optic transmitters is described. Depending on the user application, the IC is capable of driving more than 100 mA of current into a laser diode or over 50 mA into an electro-absorption or Mach-Zehnder modulator. Rise and fall times below 20 pS are achieved. The driver employs a novel dual-mode actively matched output buffer that provides a dc-coupled back termination of either 25 or 50 Ω. Compared to an output buffer with a resistive termination, this buffer dissipates only half as much power. In addition, the buffer has the the ability to reject external bias and will therefore not load bias sources used to set laser threshold currents and modulator offset voltages. The low power consumption makes the IC most suitable for co-packaging with uncooled lasers and electro-absorption modulators. The driver is fabricated in a 0.25-μm gate length production GaAs PHEMT process with substrate thru vias, thin-film resistors, and MIM capacitors     

Cheating in medical school: a survey of second-year students at 31 schools

This study examined mechanisms by which fluoxetine may reduce energy consumption and body weight. Women with binge-eating disorder (BED; n = 38) and age- and weight-matched women without BED (n = 32) monitored their dietary intake and concurrently recorded mood variables on a hand-held computer for 6 d of baseline and for 6 d after being randomly assigned to receive placebo or fluoxetine (60 mg). Fluoxetine reduced eating more than did the placebo on days 4-6 of treatment. The frequency of episodes was not affected, suggesting that fluoxetine affects satiety, not hunger. Fluoxetine did not preferentially reduce carbohydrate intake, did not affect snack consumption as compared with meal consumption, and did not affect negative-mood eating more than positive-mood eating, nor did fluoxetine affect subjects' mood ratings. Benefits of fluoxetine were of approximately equal magnitude for women with and without BED. However, women who reported higher energy consumption at baseline were more responsive to fluoxetine than were women who reported lower energy consumption at baseline, and binge-eating status was associated with greater energy consumption at all time points, including baseline. Fluoxetine affects dietary intake within 4 d of its consumption, and if future research shows that this remains true on repeated applications, this drug may be useful for short periods when difficulty with overeating is anticipated, such as during vacations.     

Baculovirus-mediated expression of Plasmodium falciparum erythrocyte binding antigen 175 polypeptides and their recognition by human antibodies

The erythrocyte binding antigen EBA-175 is a 175-kDa Plasmodium falciparum protein which mediates merozoite invasion of erythrocytes in a sialic acid-dependent manner. The purpose of this study was to produce recombinant EBA-175 polypeptide domains which have previously been identified as being involved in the interaction of EBA-175 with erythrocytes and to determine whether these polypeptides are recognized by malaria-specific antibodies. The eba-175 gene was cloned by PCR from genomic DNA isolated from the 3D7 strain of P. falciparum. The predicted protein sequence was highly conserved with that predicted from the published eba-175 gene sequences from the Camp and FCR-3 strains of P. falciparum and contained the F segment divergent region. Purified recombinant EBA-175 polypeptide fragments, expressed as glutathione S-transferase fusion proteins in insect cells by using the baculovirus system, were recognized by antibodies present in serum from a drug-cured, malaria-immune Aotus nancymai monkey. The fusion proteins were also recognized by antibodies present in sera from individuals residing in areas where malaria is endemic. In both cases the antibodies specifically recognized the EBA-175 polypeptide portion of the fusion proteins. Antibodies raised in rabbits immunized with the recombinant fusion proteins recognized parasite proteins present in schizont-infected erythrocytes. Our results suggest that these regions of the EBA-175 protein are targets for the immune response against malaria and support their further study as possible vaccine components.     

Short-term outcomes of a motivation-enhancing approach to DUI intervention

Objective

We compared a group-delivered, theory-based, motivation-enhancing program (PRIME For Life^® – PFL, n = 450) to an intervention as usual (IAU, n = 72).

Method

Individuals convicted of a substance related offense in North Carolina, typically first offense alcohol and drug-impaired driving, participated in a PFL or IAU group. We compare the interventions on program satisfaction and changes made from preintervention to postintervention, and examined the moderating effects of demographics and alcohol dependence level.

Results

When significant, findings varied in magnitude from small to medium effects. Participants in both interventions showed intentions to use statistically significantly less alcohol and drugs in the future compared to their previous use, and differences between the groups were not statistically significant. Otherwise, findings favored PFL. PFL exhibited greater benefit than IAU on understanding tolerance, perceived risk for addiction, problem recognition, and program satisfaction. Additionally, IAU perceived less risk for negative consequences postintervention than they had at preintervention. Moderation analyses showed that the between-condition findings occurred regardless of gender, age, education, and number of alcohol dependence indicators. Additionally, younger people and those with more dependence indicators – groups of particular concern – showed the greatest change.

Conclusions

Findings suggest that a motivation-enhancing approach can be effective in producing short-term change in factors that can help facilitate and sustain behavioral change. This is consistent with previous research on the use of motivational approaches, and extends such findings to suggest promise in group-based settings and with people across demographic categories and dependence levels. Future research should focus on larger studies looking at long-term behavioral change, including recidivism.     

Organizational Virtual Communities:Exploring Motivations Behind Online Panel Participation

This special thematic section of the Journal of Computer-Mediated Communication brings together nine articles that provide a rich composite of the current research in online communities. The articles cover a range of topics, methodologies, theories and practices. Indirectly they all speak to design since they aim to extend our understanding of the field. The variety shown in these articles illustrates how broad the definition is of this rapidly growing field known as 'online communities.'     

Distribution and molecular analysis of Lyme disease spirochetes, Borrelia burgdorferi, isolated from ticks throughout California

Previous studies describing the occurrence and molecular characteristics of Lyme disease spirochetes, Borrelia burgdorferi, from California have been restricted primarily to isolates obtained from the north coastal region of this large and ecologically diverse state. Our objective was to look for and examine B. burdorferi organisms isolated from Ixodes pacificus ticks collected from numerous regions spanning most parts of California where this tick is found. Thirty-one isolates of B. burgdorferi were examined from individual or pooled I. pacificus ticks collected from 25 counties throughout the state. One isolate was obtained from ticks collected at Wawona Campground in Yosemite National Park, documenting the occurrence of the Lyme disease spirochete in an area of intensive human recreational use. One isolate from an Ixodes neotomae tick from an additional county was also examined. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblot analysis, agarose gel electrophoresis, Southern blot analysis, and the polymerase chain reaction were used to examine the molecular and genetic determinants of these uncloned, low-passage-number isolates. All of the isolates were identified as B. burgdorferi by their protein profiles and reactivities with monoclonal and polyclonal antibodies, and all the isolates were typed by the polymerase chain reaction as North American-type spirochetes (B. burgdorferi sensu stricto). Although products of the ospAB locus were identified in protein analyses in all of the isolates, several isolates contained deleted forms of this locus that would result in the expression of chimeric OspA-OspB proteins. The analysis of OspC demonstrated that this protein was widely conserved among the isolates but was also quite variable in its molecular mass and the amount of it that was expressed.     

Automated design of degenerate codon libraries

Degenerate codon libraries are frequently used in protein engineering and evolution studies but are often limited to targeting a small number of positions to adequately limit the search space. To mitigate this, codon degeneracy can be limited using heuristics or previous knowledge of the targeted positions. To automate design of libraries given a set of amino acid sequences, an algorithm (LibDesign) was developed that generates a set of possible degenerate codon libraries, their resulting size, and their score relative to a user-defined scoring function. A gene library of a specified size can then be constructed that is representative of the given amino acid distribution or that includes specific sequences or combinations thereof. LibDesign provides a new tool for automated design of high-quality protein libraries that more effectively harness existing sequence-structure information derived from multiple sequence alignment or computational protein design data.