Microfluidic integration for automated targeted proteomic assays

A dearth of protein isoform-based clinical diagnostics currently hinders advances in personalized medicine. A well-organized protein biomarker validation process that includes facile measurement of protein isoforms would accelerate development of effective protein-based diagnostics. Toward scalable protein isoform analysis, we introduce a microfluidic "single-channel, multistage" immunoblotting strategy. The multistep assay performs all immunoblotting steps: separation, immobilization of resolved proteins, antibody probing of immobilized proteins, and all interim wash steps. Programmable, low-dispersion electrophoretic transport obviates the need for pumps and valves. A three-dimensional bulk photoreactive hydrogel eliminates manual blotting. In addition to simplified operation and interfacing, directed electrophoretic transport through our 3D nanoporous reactive hydrogel yields superior performance over the state-of-the-art in enhanced capture efficiency (on par with membrane electroblotting) and sparing consumption of reagents (ca. 1 ng antibody), as supported by empirical and by scaling analyses. We apply our fully integrated microfluidic assay to protein measurements of endogenous prostate specific antigen isoforms in (i) minimally processed human prostate cancer cell lysate (1.1 pg limit of detection) and (ii) crude sera from metastatic prostate cancer patients. The single-instrument functionality establishes a scalable microfluidic framework for high-throughput targeted proteomics, as is relevant to personalized medicine through robust protein biomarker verification, systematic characterization of new antibody probes for functional proteomics, and, more broadly, to characterization of human biospecimen repositories.     

Immediate antiviral therapy appears to restrict resting CD4+ cell HIV-1 infection without accelerating the decay of latent infection

HIV type 1 (HIV-1) persists within resting CD4(+) T cells despite antiretroviral therapy (ART). To better understand the kinetics by which resting cell infection (RCI) is established, we developed a mathematical model that accurately predicts (r = 0.65, P = 2.5 × 10(-4)) the initial frequency of RCI measured about 1 year postinfection, based on the time of ART initiation and the dynamic changes in viremia and CD4(+) T cells. In the largest cohort of patients treated during acute seronegative HIV infection (AHI) in whom RCI has been stringently quantified, we found that early ART reduced the generation of latently infected cells. Although RCI declined after the first year of ART in most acutely infected patients, there was a striking absence of decline when initial RCI frequency was less than 0.5 per million. Notably, low-level viremia was observed more frequently as RCI increased. Together these observations suggest that (i) the degree of RCI is directly related to the availability of CD4(+) T cells susceptible to HIV, whether viremia is controlled by the immune response and/or ART; and (ii) that two pools of infected resting CD4(+) T cells exist, namely, less stable cells, observable in patients in whom viremia is not well controlled in early infection, and extremely stable cells that are established despite early ART. These findings reinforce and extend the concept that new approaches will be needed to eradicate HIV infection, and, in particular, highlight the need to target the extremely small but universal, long-lived latent reservoir.     

Site-Dependent Effects of tDCS Uncover Dissociations in the Communication Network Underlying the Processing of Visual Search

BackgroundThe right posterior parietal cortex (rPPC) and the right frontal eye field (rFEF) form part of a network of brain areas involved in orienting spatial attention. Previous studies using transcranial magnetic stimulation (TMS) have demonstrated that both areas are critically involved in the processing of conjunction visual search tasks, since stimulation of these sites disrupts performance.ObjectiveThis study investigated the effects of long term neuronal modulation to rPPC and rFEF using transcranial direct current stimulation (tDCS) with the aim of uncovering sharing of these resources in the processing of conjunction visual search tasks.MethodsParticipants completed four blocks of conjunction search trials over the course of 45 min. Following the first block they received 15 min of either cathodal or anodal stimulation to rPPC or rFEF, or sham stimulation.ResultsA significant interaction between block and stimulation condition was found, indicating that tDCS caused different effects according to the site (rPPC or rFEF) and type of stimulation (cathodal, anodal, or sham). Practice resulted in a significant reduction in reaction time across the four blocks in all conditions except when cathodal tDCS was applied to rPPC.ConclusionsThe effects of cathodal tDCS over rPPC are subtler than those seen with TMS, and no effect of tDCS was evident at rFEF. This suggests that rFEF has a more transient role than rPPC in the processing of conjunction visual search and is robust to longer-term methods of neuro-disruption. Our results may be explained within the framework of functional connectivity between these, and other, areas.     

The effect of Er:YAG laser irradiation on hydroxyapatite-coated implants and fluoride-modified TiO2-blasted implant surfaces: a microstructural analysis

The purpose of this study was to evaluate the microscopic changes and surface roughness on hydroxyapatite (HA)-coated implants following exposure to different powers and durations of Er:YAG laser irradiation in order to determine the proper pulse energy level and irradiation time. Ten HA-coated implants and ten fluoride-modified TiO₂ implants were used. The implants were divided into a control (one implant) and test group (nine implants) for each implant type. Implants in the test groups were sub-divided into three groups (three implants per group) based on the applied laser pulse energy and irradiation time. The measurement of surface roughness was performed on all implants in the test groups using a white light interferometer before and after laser irradiation. R _a values were recorded and compared in order to evaluate changes in surface roughness. For HA-coated implants, the R _a values increased in all test groups after laser irradiation. However, mean R _a values in the fluoride-modified TiO₂-blasted implant test group were decreased after irradiation. There was no statistical difference. Scanning electron microscope analysis revealed surface alterations in both the HA-coated and fluoridated TiO₂-blasted implants irradiated for 1.5 min at 100 mJ/pulse, 10 Hz. When the pulse energy and irradiation time increased, greater surface alterations, including surface flattening and microfractures, were observed. In conclusion, the results of the current study suggest that no changes could be observed in both HA-coated implants and fluoride-modified TiO₂-blasted implants after irradiation at an intensity of 100 mJ/pulse, 10 Hz for 1 min performed to achieve surface detoxification.     

Bronchodilators in Heart Failure Patients With COPD: Is It Time for a Clinical Trial?

Chronic heart failure (HF) and chronic obstructive pulmonary disease (COPD) commonly coexist, and patients with both diseases fare worse than those with either disease alone. Several factors may contribute to worse outcomes, including an increased burden of care related to greater disease complexity, an overlap of symptoms resulting in misapplication of therapy, and the adverse effects of treatment for one disease on the symptoms and outcomes related to the other. For example, there are conflicting data about the cardiovascular risks of bronchodilators in HF patients who may experience worse outcomes with inhaled beta-2 agonists via arrhythmogenesis, ischemia, and/or attenuation of beta-blocker benefits. In contrast, the long-acting anticholinergic class of bronchodilators has a more reassuring safety profile. Anticholinergic bronchodilators may be the preferred first-line agents for COPD patients with comorbid HF, yet data supporting these recommendations are limited. Therapeutic trials in COPD patients have generally excluded patients with significant HF and vice-versa. This paper reviews bronchodilator therapy in HF and proposes a randomized trial designed to enroll patients with significant COPD and HF to determine the risks and/or benefits of adding a long-acting beta-2 agonist to patients currently taking a long-acting anticholinergic agent.     

Age identity, self-rated health, and life satisfaction among older adults in Dakar, Senegal

The objectives of this quantitative study were to (1) ascertain to what extent older adults aged 50 and above feel and desire to be younger than their age, and classify themselves as young versus old; (2) compare these patterns with those found among other cross-cultural populations; and (3) assess the extent to which self-rated health and life satisfaction predict age identities. This study was carried out on a sample of 500 dwellers of the Senegalese capital aged 50 and older. This sample was constructed using the quota method to strive for representativeness. Most of the respondents wanted to be younger than their chronological age (51.8 %), but only 27.8 % felt younger than they were. Moreover, 80 % of the sample claimed to be old. Self-rated health predicted felt age and the feeling of being old. Furthermore, the less-satisfied Dakar residents were with their life, the younger they wanted to be. We first discuss our results in a comparative perspective focused on how orientations toward individualism and collectivism could be related to age identity, and on demographic characteristics of the Senegalese population—where life expectancy is 59.3 years old. We then analyze the relevance of age identity dimensions as indicators of successful aging in Dakar.