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.     

Iodine Deficiency and the Brain: Effects and Mechanisms

Iodine is an essential micronutrient needed in human diets. As iodine is an integral component of thyroid hormone, it mediates the effects of thyroid hormone on brain development. Iodine deficiency is the most prevalent and preventable cause of mental impairment in the world. The exact mechanism through which iodine influences the brain is unclear, but is generally thought to begin with genetic expression. Many brain structures and systems appear to be affected with iodine deficiency, including areas such as the hippocampus, microstructures such as myelin, and neurotransmitters. The clearest evidence comes from the studies examining cognition in the cases of iodine deprivation or interventions involving iodine supplementation. Nevertheless, there are many inconsistencies and gaps in the literature of iodine deficiency, especially over the lifespan. This paper summarizes the literature on this topic, suggests a causal mechanism for iodine's effect on the brain, and indicates areas for the future research (e.g., using magnetic resonance imaging (MRI) and functional MRI to examine how iodine supplementation facilitates cognitive functioning).     

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.     

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)     

When the face reveals what words do not: Facial expressions of emotion, smiling, and the willingness to disclose childhood sexual abuse.

For survivors of childhood sexual abuse (CSA), verbal disclosure is often complex and painful. The authors examined the voluntary disclosure-nondisclosure of CSA in relation to nonverbal expressions of emotion in the face. Consistent with hypotheses derived from recent theorizing about the moral nature of emotion, CSA survivors who did not voluntarily disclose CSA showed greater facial expressions of shame, whereas CSA survivors who voluntarily disclosed CSA expressed greater disgust. Expressions of disgust also signaled sexual abuse accompanied by violence. Consistent with recent theorizing about smiling behavior, CSA nondisclosers made more polite smiles, whereas nonabused participants expressed greater genuine positive emotion. Discussion addressed the implications of these findings for the study of disclosure of traumatic events, facial expression, and the links between morality and emotion. (PsycINFO Database Record (c) 2010 APA, all rights reserved)