Multiplex quantitative competitive polymerase chain reaction based on a multianalyte hybridization assay performed on spectrally encoded microspheres

Quantitative polymerase chain reaction (PCR) may be performed by two general approaches, namely, real-time PCR and quantitative competitive PCR (QC-PCR). QC-PCR makes use of the concept of a DNA competitor, which is the "gold standard" approach to circumvent the problem of the variation of amplification efficiency. However, QC-PCR in its classical form is a low-throughput method since it requires titration of each sample with the competitor followed by electrophoresis. The throughput of QC-PCR has been improved by capillary electrophoresis and microtiter well-based hybridization assays. The present work introduces a multiplex QC-PCR method, which is based on a multianalyte hybridization assay that is performed on spectrally encoded microspheres. The DNA competitors use the same primers and have equal size with their corresponding target DNA sequences but differ in a short (24 bp) centrally located sequence. Following multiplex PCR, biotinylated amplification products from all DNA targets and competitors are heat-denatured and hybridized with oligonucleotide probes, which are attached to addressable sets of fluorescent microspheres. The hybrids react with a streptavidin-phycoerythrin conjugate. The microspheres are then analyzed by flow cytometry employing two lasers. A red laser line is used for classification of the microspheres, and a green line excites phycoerythrin, whose fluorescence is related to the concentration of the analyte DNA. As a model, we have developed a multiplex quantitative competitive PCR assay for four targets. The amplification products from targets and competitors (a total of 8 DNA fragments) are determined simultaneously by the multianalyte hybridization assay. The limits of quantification for the hybridization assay of all amplified DNA fragments are below 13 pM. The multiplex quantitative competitive PCR assay detects approximately 500 copies from each target DNA. To our knowledge, the proposed method is the only approach to quantitative PCR that offers such a high potential for multiplexing.     

The nature and structure of correlations among Big Five ratings: The halo-alpha-beta model.

In light of consistently observed correlations among Big Five ratings, the authors developed and tested a model that combined E. L. Thorndike’s (1920) general evaluative bias (halo) model and J. M. Digman’s (1997) higher order personality factors (alpha and beta) model. With 4 multitrait–multimethod analyses, Study 1 revealed moderate convergent validity for alpha and beta across raters, whereas halo was mainly a unique factor for each rater. In Study 2, the authors showed that the halo factor was highly correlated with a validated measure of evaluative biases in self-ratings. Study 3 showed that halo is more strongly correlated with self-ratings of self-esteem than self-ratings of the Big Five, which suggests that halo is not a mere rating bias but actually reflects overly positive self-evaluations. Finally, Study 4 demonstrated that the halo bias in Big Five ratings is stable over short retest intervals. Taken together, the results suggest that the halo-alpa-beta model integrates the main findings in structural analyses of Big Five correlations. Accordingly, halo bias in self-ratings is a reliable and stable bias in individuals’ perceptions of their own attributes. Implications of the present findings for the assessment of Big Five personality traits in monomethod studies are discussed. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Unraveling the Role of Histone Variant CENP-A and Chaperone HJURP Expression in Thymic Epithelial Neoplasms

Background: Recent advances demonstrate the role of chromatin regulators, including histone variants and histone chaperones, in cancer initiation and progression. Methods: Histone H3K4me3, histone variant centromere protein (CENP-A) and histone chaperones Holliday junction recognition protein (HJURP) as well as DAXX expression were examined immunohistochemically in 95 thymic epithelial tumor (TET) specimens. Our results were compared with the expression profile of DAXX, HJURP and CENP-A in gene expression profiling interactive analysis (GEPIA2). Results: The lymphocyte-poor B3- and C-type TETs were more frequently DAXX negative (p = 0.043). B3 and C-Type TETs showed higher cytoplasmic and nuclear CENP-A (p = 0.007 and p = 0.002) and higher cytoplasmic HJURP H-score (p < 0.001). Higher nuclear CENP-A and cytoplasmic HJURP expression was associated with advanced Masaoka–Koga stage (p = 0.048 and p < 0.001). A positive correlation between HJURP and CENP-A was also observed. The presence of cytoplasmic CENP-A expression was correlated with a favorable overall survival (p = 0.03). CENP-A overexpression in survival analysis of TCGA TETs showed similar results. H3K4me3 expression was not associated with any clinicopathological parameters. Conclusions: Our results suggest a significant interaction between CENP-A and HJURP in TETs. Moreover, we confirmed the presence of a cytoplasmic CENP-A immunolocalization, suggesting also a possible favorable prognostic value of this specific immunostaining pattern.     

Flavour Qualities of Frozen Sweetcorn are Affected by Genotype and Blanching

Peroxidase activity is used by the food processing industry as an indicator of adequacy of the blanching process and to predict off-flavour development in frozen food. Research reported herein showed that total peroxidase activity did not parallel flavour changes in frozen unblanched supersweet (sh2) or sugary enhanced (su1/se) sweet corn genotypes. Frozen corn-on-the-cob of ‘Florida Staysweet’ (sh2), ‘Merit’ (standard sweet—su1) and ‘Bodacious’ (su1/se), blanched and unblanched, was subjected to sensory evaluation and peroxidase analysis following frozen storage up to 12 months. Trained taste panellists rated the unblanched sh2 and su1/se corn as acceptable up to 8 months of frozen storage. Kernels were cut from cobs after 0 and 12 months of storage. Proteins were extracted from acetone powders of kernels and separated by isoelectric focusing and native PAGE. Banding patterns differed according to genotype and storage duration. These results suggested molecular differences in peroxidase isozymes among the sweet corn genotypes which could be involved in off-flavour development. All genotypes contained a peroxidase isozyme having a molecular mass of 80 kDa and pI of 4·5. The su1/se and sh2 genotypes produced an additional peroxidase band of 13·8 kDa. An additional peroxidase isozyme (pI 5·4) appeared in extracts from the su1 genotype after 12 months storage. Although changes in total peroxidase activity may not predict flavour changes in all genotypes, the presence or absence of certain peroxidase isozymes may be useful in predicting off-flavour development in su1 frozen corn.     

Nanostructured Poly(styrene‐b‐butadiene‐b‐styrene) (SBS) Membranes for the Separation of Nitrogen from Natural Gas

The preparation and characterization of new, tailor‐made polymeric membranes using poly(styrene‐b‐butadiene‐b‐styrene) (SBS) triblock copolymers for gas separation are reported. Structural differences in the copolymer membranes, obtained by manipulation of the self‐assembly of the block copolymers in solution, are characterized using atomic force microscopy, transmission electron microscopy, and the transport properties of three gases (CO₂, N₂, and CH₄). The CH₄/N₂ ideal selectivity of 7.2, the highest value ever reported for block copolymers, with CH₄ permeability of 41 Barrer, is obtained with a membrane containing the higher amount of polybutadiene (79 wt%) and characterized by a hexagonal array of columnar polystyrene cylinders normal to the membrane surface. Membranes with such a high separation factor are able to ease the exploitation of natural gas with high N₂ content. The CO₂/N₂ ideal selectivity of 50, coupled with a CO₂ permeability of 289 Barrer, makes SBS a good candidate for the preparation of membranes for the post‐combustion capture of carbon dioxide.     

Absolute quantification of the alleles in somatic point mutations by bioluminometric methods based on competitive polymerase chain reaction in the presence of a locked nucleic acid blocker or an allele-specific primer

In somatic (acquired) point mutations, the challenge is to quantify minute amounts of the mutant allele in the presence of a large excess of the normal allele that differs only in a single base pair. We report two bioluminometric methods that enable absolute quantification of the alleles. The first method exploits the ability of a locked nucleic acid (LNA) oligonucleotide to bind to and inhibit effectively the polymerase chain reaction (PCR) amplification of the normal allele while the amplification of the mutant allele remains unaffected. The second method employs allele-specific PCR primers, thereby allowing the amplification of the corresponding allele only. DNA internal standards (competitors) are added to the PCR mixture to compensate for any sample-to-sample variation in the amplification efficiency. The amplification products from the two alleles and the internal standards are quantified by a microtiter well-based bioluminometric hybridization assay using the photoprotein aequorin as a reporter. The methods allow absolute quantification of less than 300 copies of the mutant allele even in samples containing less than 1% of the mutant allele.     

Some Dared Call It Torture: Cultural Resonance,Abu Ghraib,and a Selectively Echoing Press

This study draws upon research on “indexing” and “cascading activation” to explore U.S. political and news discourse surrounding the Abu Ghraib prison scandal. Specifically, we systematically analyze White House, military, congressional, and news messages. In so doing, we incorporate scholarship on social identity theory to suggest why news media challenge certain White House frames but uncritically echo others. Our data demonstrate that White House frames were consistently challenged by Democrats in the opposing party, but that these competing congressional messages were largely absent in news coverage. These results challenge previous research on news coverage of Abu Ghraib. We discuss how these patterns align with and expand Entman's cascading activation model of press‐state relations, and consider the implications for future scholarship.     

Feeling Better About Doing Worse: Social Comparisons Within Romantic Relationships.

The authors examined the role of closeness between self and partner in determining the impact of social comparisons within intimate relationships. To the extent that one's partner is a central aspect of one's identity, one may be able to restore one's positive self-regard following an upward comparison with the partner by turning to the relationship as a self-affirmational resource. Studies 1 and 2 examined reactions to imagined comparisons; Studies 3 and 4 examined reactions to actual comparison feedback. Across studies, closeness moderated the impact of upward comparisons with the partner; that is, higher closeness participants responded to a more successful partner by focusing on their relationship-related strengths. However, closeness did not moderate the impact of downward comparisons with the partner. (PsycINFO Database Record (c) 2010 APA, all rights reserved)