Universal pre-mixing dry-film stickers capable of retrofitting existing microfluidics.

Integrating microfluidic mixers into lab-on-a-chip devices remains challenging yet important for numerous applications including dilutions, extractions, addition of reagents or drugs, and particle synthesis. High-efficiency mixers utilize large or intricate geometries that are difficult to manufacture and co-implement with lab-on-a-chip processes, leading to cumbersome two-chip solutions. We present a universal dry-film microfluidic mixing sticker that can retrofit pre-existing microfluidics and maintain high mixing performance over a range of Reynolds numbers and input mixing ratios. To attach our pre-mixing sticker module, remove the backing material and press the sticker onto an existing microfluidic/substrate. Our innovation centers around the multilayer use of laser-cut commercially available silicone-adhesive-coated polymer sheets as microfluidic layers to create geometrically complex, easy to assemble designs that can be adhered to a variety of surfaces, namely, existing microfluidic devices. Our approach enabled us to assemble the traditional yet difficult to manufacture "F-mixer" in minutes and conceptually extend this design to create a novel space-saving spiral F-mixer. Computational fluid dynamic simulations and experimental results confirmed that both designs maintained high performance for 0.1 < Re < 10 and disparate input mixing ratios of 1:10. We tested the integration of our system by using the pre-mixer to fluorescently tag proteins encapsulated in an existing microfluidic. When integrated with another microfluidic, our pre-mixing sticker successfully combined primary and secondary antibodies to fluorescently tag micropatterned proteins with high spatial uniformity, unlike a traditional pre-mixing "T-mixer" sticker. Given the ease of this technology, we anticipate numerous applications for point-of-care devices, microphysiological-systems-on-a-chip, and microfluidic-based biomedical research.

War-related stressors are associated with asthma risk among older Kuwaitis following the 1990 Iraqi invasion and occupation.

BACKGROUND: Contemporary warfare involving civilian populations is a growing public health concern. In addition to the psychological impact, war-related trauma may result in physiological alterations and even broader health effects. Associations were examined between war-related stressors and incident asthma in elderly Kuwaiti civilians following the Iraqi invasion. METHODS: A random sample of all Kuwaiti nationals aged 50-69 years on the day prior to the invasion were identified. Among the 7873 meeting eligibility criteria, 5567 (71%) agreed to participate and 5028 completed the questionnaire (91% of those eligible). Of these, 3759 were in Kuwait during the invasion, of whom 2294 were alive at follow-up. After exclusions for prevalent asthma or missingness on covariates, 2066 were available for analysis. War-related experiences were summarised into a continuous score using Rasch modelling. Relative Cox proportional hazard rates (HR) were calculated for asthma adjusting for covariates. RESULTS: Over 13 years of follow-up, physician-diagnosed asthma was reported by 66/996 (6.6%) men and 104/1070 (9.7%) women. In models adjusted for gender, socioeconomic status, smoking, BMI, and air pollution related to burning oil fires, those reporting highest stress exposure were more than twice as likely to report asthma (HR 2.3, 95% CI 1.3, 3.9) compared to civilians reporting no stressors. Experiences were more salient when anchored to fear for loss of life. CONCLUSIONS: War-related trauma is associated with increased asthma risk in these elderly civilians. Although prior research has documented the significant and persistent psychological toll of war, these findings implicate even broader health effects.

New York 1 and Sin Nombre viruses are serotypically distinct viruses associated with hantavirus pulmonary syndrome.

New York 1 virus (NY-1) and Sin Nombre virus (SN) are associated with hantavirus pulmonary syndrome (HPS). NY-1 and SN are derived from unique mammalian hosts and geographic locations but have similar G1 and G2 surface proteins (93 and 97% identical, respectively). Focus reduction neutralization assays were used to define the serotypic relationship between NY-1 and SN. Sera from NY-1-positive Peromyscus leucopus neutralized NY-1 and SN at titers of >/=1/3,200 and 16-fold-lower neutralizing titers to NY-1 than to SN. Reference sera to Hantaan, Seoul, and Prospect Hill viruses also failed to neutralize NY-1. These results indicate that SN and NY-1 define unique hantavirus serotypes and implicate the presence of additional HPS-associated hantavirus serotypes in the Americas.

Autoradiographic identification of a gastrin receptor on the human parietal cell.

Gastrin plays an important role in regulating gastric acid secretion and gastrointestinal mucosal growth but its cellular sites of action in man have not been determined. Using cryostat sections of gastric mucosal tissue we have identified (125I-gastrin binding followed by fixation-wet emulsion autoradiography) and characterized (125I-gastrin binding followed by counting) a gastrin receptor binding site in the human stomach. This site displayed binding characteristics similar to those observed in isolated cell systems: specifically, 125I-gastrin binding was rapid (t1/2 approximately 10 min at 37 degrees C), temperature-dependent (3.5 fold more radioligand bound at 22 degrees C than at 4 degrees C) and saturable. The binding of the radioligand was also tissue specific and was five-fold greater in the gastric body than in the gastric antrum and duodenum. In the autoradiographs, silver grains were localized only to parietal cells and not to other epithelial cell types. In the presence of 40 nM gastrin grains were no longer present over parietal cells demonstrating that these sites were both saturable and of high affinity. These data provide the first demonstration of gastrin binding sites (putative receptors) on parietal cells in the human stomach and suggest that gastrin acts directly on these cells to help regulate gastric acid secretion and/or mucosal growth.

Cloning sequencing and expression of the gene encoding the VP2 protein of the human group B rotavirus, ADRV.

Adult diarrheal rotavirus, ADRV, is a noncultivable human group B rotavirus. A complete cDNA copy of ADRV gene segment 2 has been cloned and sequenced. Gene segment 2 contains 2844 bases and encodes one long open reading frame beginning at base 14 and terminating at base 2812. Gene 2 encodes a protein containing 933 amino acids with a calculated molecular weight of 105.6 kDa and a pl of 5.5. The gene 2 polypeptide contains significant homology with the VP2 protein which comprises the core of group A rotavirus strains. The gene 2 protein has been expressed in a rabbit reticulocyte lysate in vitro and is identical in molecular mass with a protein previously demonstrated to be present on iodinated EDTA-treated virions and from in vitro translations of total ADRV mRNA. A recombinant baculovirus containing gene segment 2 has been constructed and used to express the encoded VP2 equivalent protein. The expressed VP2 protein is immunoprecipitable by hyperimmune anti-ADRV serum, porcine group B infection serum, and human convalescent serum but not by hyperimmune serum to group A rotavirus. Our results suggest that ADRV gene segment 2 encodes the VP2 protein equivalent to group A rotavirus strains present in the core of the group B virion.

An association between air pollution and mortality in six U.S. cities.

BACKGROUND: Recent studies have reported associations between particulate air pollution and daily mortality rates. Population-based, cross-sectional studies of metropolitan areas in the United States have also found associations between particulate air pollution and annual mortality rates, but these studies have been criticized, in part because they did not directly control for cigarette smoking and other health risks. METHODS: In this prospective cohort study, we estimated the effects of air pollution on mortality, while controlling for individual risk factors. Survival analysis, including Cox proportional-hazards regression modeling, was conducted with data from a 14-to-16-year mortality follow-up of 8111 adults in six U.S. cities. RESULTS: Mortality rates were most strongly associated with cigarette smoking. After adjusting for smoking and other risk factors, we observed statistically significant and robust associations between air pollution and mortality. The adjusted mortality-rate ratio for the most polluted of the cities as compared with the least polluted was 1.26 (95 percent confidence interval, 1.08 to 1.47). Air pollution was positively associated with death from lung cancer and cardiopulmonary disease but not with death from other causes considered together. Mortality was most strongly associated with air pollution with fine particulates, including sulfates. CONCLUSIONS: Although the effects of other, unmeasured risk factors cannot be excluded with certainty, these results suggest that fine-particulate air pollution, or a more complex pollution mixture associated with fine particulate matter, contributes to excess mortality in certain U.S. cities.

Identification and baculovirus expression of the VP4 protein of the human group B rotavirus ADRV.

A complete cDNA copy of the fourth RNA segment of the human group B rotavirus adult diarrheal rotavirus (ADRV) has been cloned into lambda phage and excised into plasmid pSK Bluescript. Gene segment 4 contains 2,303 bases and encodes one long open reading frame beginning at base 16 and terminating at base 2263. The encoded protein contains 749 amino acids, with a calculated molecular mass of 84.4 kDa and a pI of 6.1. Gene 4 cDNA was inserted into a recombinant baculovirus via homologous recombination. The gene 4 polypeptide migrates at 84 kDa when expressed either by a recombinant baculovirus or in vitro in a rabbit reticulocyte lysate. The gene 4 protein is immunoprecipitable by hyperimmune serum to ADRV, human ADRV convalescent-phase serum, a porcine group B rotavirus infection serum, and a monoclonal antibody made to ADRV virion. Guinea pig hyperimmune serum to the baculovirus-expressed ADRV VP4 protein recognizes virus and immunoprecipitates an 84-kDa protein from in vitro translations of total ADRV mRNA. In addition, the gene 4-encoded protein shares significant amino acid identity and similarity with the group A rotavirus VP4 protein. This information, together with our previous identification of an 84-kDa protein present on iodinated intact virion but not EDTA-treated ADRV, suggests that gene 4 encodes the VP4 protein equivalent present on the outer capsid of ADRV. The ADRV VP4 protein is also 58% identical to the IDIR rat group B rotavirus gene segment 3 protein. The substantial differences between these two group B VP4 proteins suggests that they are distantly related and likely to define two different group B rotavirus VP4 serotypes. The baculovirus-expressed VP4 protein should be useful for developing serotyping reagents and tests for human and animal group B rotaviruses as well as for addressing the role of VP4 in ADRV neutralization.

Baculovirus expression of the ADRV gene 5 encoded protein produces an oligomerized, antigenic, and immunogenic VP6 protein.

Adult diarrheal rotavirus (ADRV) is a currently noncultivatable group B human rotavirus responsible for epidemic outbreaks of gastroenteritis in China. Gene segment 5 of ADRV encodes the major inner capsid protein, VP6. ADRV gene 5 was inserted into a recombinant baculovirus by homologous recombination between baculovirus shuttle plasmid pACYM1-AD5 and AcNPV genomic DNA. Baculovirus recombinants were selected visually and plaque purified and VP6 expression was detected by Coomassie staining of PAGE-separated proteins. The baculovirus-expressed gene 5 polypeptide is 44 kDa, the same as for the major inner capsid protein present on EDTA-treated ADRV virions and in vitro-expressed VP6 protein. The expressed protein is oligomeric and in the absence of reducing agents multimerizes to apparent trimer, hexamer, and greater molecular mass as assayed by SDS-PAGE. The VP6 protein is immunoprecipitable by hyperimmune serum to ADRV, human ADRV convalescent serum, by a group B-specific monoclonal antibody and by porcine group B rotavirus infection serum. The baculovirus-expressed protein is immunogenic and antibodies to the expressed protein recognize ADRV virions. The ADRV VP6 protein should be useful for developing diagnostic assays for serum antibodies to group B rotavirus as well as for generating hyperimmune serum and monoclonal antibodies for detecting viral antigen from ADRV and other group B rotaviruses.

Pulmonary function between 6 and 18 years of age.

Pulmonary function of children aged 6-18 years is described based on 82,462 annual measurements of forced vital capacity (FVC), forced expired volume in 1 second (FEV1), and forced expiratory flow between 25% and 75% of FVC (FEF25-75%) from 11,630 white children and 989 black children. Median height, FVC, FEV1, FEV1/FVC, and FEF25-75% for each 3 months of age are compared among race and sex subgroups. Race- and sex-specific percentile distributions of FVC, FEV1, FEV1/FVC, and FEF25-75% are presented for each centimeter of height (growth curves). For the same height, boys have greater lung function values than girls, and whites have greater ones than blacks. Lung function increases linearly with age until the adolescent growth spurt at about age 10 years in girls and 12 in boys. The pulmonary function vs. height relationship shifts with age during adolescence. Thus, a single equation or the pulmonary function-height growth chart alone does not completely describe growth during the complex adolescent period. Nevertheless, race- and sex-specific growth curves of pulmonary function vs. height make it easy to display and evaluate repeated measures of pulmonary function for an individual child. Race-, sex-, and age-specific regression equations based on height are provided, which permit the evaluation of growth during adolescence with improved accuracy and, more importantly, in comparison with previous observations for the same child.

Chronic obstructive pulmonary disease mortality in six U.S. cities.

Results are presented from a 9- to 12-yr mortality follow-up of 8,427 white adults in six U.S. cities between 25 and 74 yr of age at enrollment. Survival analyses were performed for all causes on 941 confirmed deaths, and for specific primary causes for the subset of 851 death with death certificates. Relative level of FEV1 compared with predicted was a strong predictor of sex-specific chronic obstructive pulmonary disease (COPD), cardiovascular (CV), and all-cause mortality, even after adjusting for age, respiratory symptoms, and smoking. Even in this relatively large cohort, the total number of respiratory deaths was small, and no trend in COPD mortality could be determined over the period of study.

The association between health status and the performance of excessively variable spirometry tests in a population-based study in six U.S. cities.

The relationship between 6 chronic respiratory symptoms and the performance of an excessively variable FEV1 (test failure) was examined among 8,522 white adults in 6 U.S. cities. A total of 747 (8.9%) performed an excessively variable FEV1 according to the American Thoracic Society criterion. After adjusting for smoking, age, and city of residence in 6 separate logistic regression models, the odds ratios for FEV1 failure among men were 2.32, 1.39, 1.40, 1.82, 2.61, 1.92 for moderate breathlessness, chronic cough, phlegm, wheeze, asthma, and recurrent chest illness, respectively. Among women, FEV1 failure was significantly associated with moderate breathlessness, chronic phlegm, wheeze, and asthma with odds ratios of 1.55, 1.45, 1.62, and 1.95, respectively. When all symptoms were evaluated simultaneously in a single logistic regression model, only breathlessness and asthma remained associated with FEV1 failure; odds ratio = 1.97 for asthma and 2.03 for breathlessness among men and 1.53 for both asthma and breathlessness among women. The 11-yr mortality experience of subjects with test failure, as defined by 2 different criteria, was compared to that of the quartile of the cohort with the highest cross-sectional test results. After adjusting for age, gender, and smoking, the relative risks of mortality were 1.62 and 1.98 for subjects with an FEV1 failure as defined by the ATS and 6-Cities criteria, respectively, and 1.99 and 1.90 for the groups with FVC failure as defined by the 2 criteria. Thus test failure is almost as strong a predictor of mortality as poor FEV1.

Distribution of forced expiratory volume in one second and forced vital capacity in healthy, white, adult never-smokers in six U.S. cities.

As part of a longitudinal study of the respiratory health effects of air pollution, we measured the lung function of 2,454 white adults 25 to 74 yr of age who had never smoked and who reported no respiratory symptoms. These measurements were analyzed to develop a simple model for the cross-sectional dependence of pulmonary function on height, sex, and age. Both forced expiratory volume in one second (FEV1) and forced vital capacity (FVC) can be effectively standardized for body size by dividing each pulmonary function measurement by the square of the standing height (HT2). The age-specific distribution of these standardized measurements is approximately Gaussian, with variance that is independent of age. Plots of FEV1/HT2 and FVC/HT2 against age showed a nonlinear relationship consistent with an increase in the rate of pulmonary function loss with age. On the basis of these graphic analyses, both pulmonary function measurements were fitted to a four-parameter normative model including sex and linear and quadratic terms in age as dependent variables. This model gave predictions that were very close to those from more complicated models currently in use. Predicted percentile levels were calculated for each sex and age, and shown to describe the observations well. The estimated annual change in height-standardized lung function based on the cross-sectional model was compared with the observed change between the first and second examinations of these adults 3 yr later. The observed changes were close to predicted values, except for subjects younger than 35 yr of age at their first examination. The observed change was larger for men than for women. Such simple longitudinal comparisons are subject to selection bias. In this study, subjects in the lowest quartile of FEV1/HT2 for their age and sex at the first examination had a lower probability of providing a lung function measurement 3 yr later.