A Dynamic Data Correction Method for Enhancing Resolving Power of Integrated Spectra in Spectroscopic Analysis.

A dynamic data correction method embedded in the process of data acquisition improves spectral quality. The method minimizes the impact of random errors in spectroscopic measurements by correcting peak positions in every single-scan spectrum. The method is fast enough to facilitate online data correction. The integration of corrected spectra improves resolving power and signal-to-noise ratio. The correction method can apply to most analytical spectra. In mass spectrometry and Raman spectroscopy, observations show that it improved the average resolving power by roughly 40-150% and revealed unresolved spectral features.

An Efficient Sample Preparation Method to Enhance Carbohydrate Ion Signals in Matrix-assisted Laser Desorption/Ionization Mass Spectrometry.

Sample preparation is a critical process in mass spectrometry (MS) analysis of carbohydrates. Although matrix-assisted laser desorption/ionization (MALDI) MS is the method of choice in carbohydrate analysis, poor ion signal and data reproducibility of carbohydrate samples continue to be severe problems. For quantitative analysis of carbohydrates, an effective analytical protocol providing superior data quality is necessary. This video demonstrates sample preparation protocols to improve signal intensity and minimize data variation of carbohydrates in MALDI-MS. After drying and crystallization of sample droplets, the crystal morphology is reformed by methanol before mass spectrometric analysis. The enhancement in carbohydrate signal is examined with MALDI imaging mass spectrometry (IMS). Experimental results show that crystal reformation adjusts crystalline structures and redistributes carbohydrate analytes. In comparison with the dried droplet preparation method in conventional MALDI-MS, reforming carbohydrate crystal morphologies with methanol shows significantly better signal intensity, ion image distribution, and data stability. Since the protocols demonstrated herein do not involve changes in sample composition, they are generally applicable to various carbohydrates and matrixes.

Enhancing carbohydrate ion yield by controlling crystalline structures in matrix-assisted laser desorption/ionization mass spectrometry.

Carbohydrate analysis is challenging due to lack of sensitive detection and efficient separation methods. Although matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) is a sensitive tool, the low ionization efficiency of carbohydrates makes mass analyses inefficient. This work systematically examines the correlation between MALDI-MS sensitivity and carbohydrate sample morphology. Depending on the properties of the matrix used, the morphology changes through sample recrystallization after drying or imposition of hydrodynamic flows during droplet drying. Observation shows that amorphous solids and finer crystals offer higher carbohydrate sensitivity and spatial homogeneity than larger crystals. Clear evidences of an inverse correlation between sensitivity and crystal size are obtained when various kinds of carbohydrates are mixed with different matrixes. Similar experiments on proteins and peptides showed a negative or negligible effect. The result serves as a general guideline for improving efficiency in routine carbohydrate analysis.

PPL2ab neurons restore sexual responses in aged Drosophila males through dopamine.

Male sexual desire typically declines with ageing. However, our understanding of the neurobiological basis for this phenomenon is limited by our knowledge of the brain circuitry and neuronal pathways controlling male sexual desire. A number of studies across species suggest that dopamine (DA) affects sexual desire. Here we use genetic tools and behavioural assays to identify a novel subset of DA neurons that regulate age-associated male courtship activity in Drosophila. We find that increasing DA levels in a subset of cells in the PPL2ab neuronal cluster is necessary and sufficient for increased sustained courtship in both young and aged male flies. Our results indicate that preventing the age-related decline in DA levels in PPL2ab neurons alleviates diminished courtship behaviours in male Drosophila. These results may provide the foundation for deciphering the circuitry involved in sexual motivation in the male Drosophila brain.

Dengue virus enhances thrombomodulin and ICAM-1 expression through the macrophage migration inhibitory factor induction of the MAPK and PI3K signaling pathways.

Dengue virus (DV) infections cause mild dengue fever (DF) or severe life-threatening dengue hemorrhagic fever (DHF). The mechanisms that cause hemorrhage in DV infections remain poorly understood. Thrombomodulin (TM) is a glycoprotein expressed on the surface of vascular endothelial cells that plays an important role in the thrombin-mediated activation of protein C. Prior studies have shown that the serum levels of soluble TM (sTM) and macrophage migration inhibitory factor (MIF) are significantly increased in DHF patients compared to levels in DF patients or normal controls. In this study, we investigated how MIF and sTM concentrations are enhanced in the plasma of DHF patients and the potential effect of MIF on coagulation through its influence on two factors: thrombomodulin (TM) and intercellular adhesion molecule-1 (ICAM-1) in endothelial cells and monocytes. Recombinant human macrophage migration inhibitory factor (rMIF) was used to treat monocytic THP-1 cells and endothelial HMEC-1 cells or primary HUVEC cells. The subsequent expression of TM and ICAM-1 was assessed by immunofluorescent staining and flow cytometry analysis. Additionally, the co-incubation of THP-1 cells with various cell signaling pathway inhibitors was used to determine the pathways through which MIF mediated its effect. The data provided evidence that severe DV infections induce MIF expression, which in turn stimulates monocytes or endothelial cells to express TM and ICAM-1 via the Erk, JNK MAPK and the PI3K signaling pathways, supporting the idea that MIF may play an important role as a regulator of coagulation.

A hormone receptor-based transactivator bridges different binary systems to precisely control spatial-temporal gene expression in Drosophila.

The GAL4/UAS gene expression system is a precise means of targeted gene expression employed to study biological phenomena in Drosophila. A modified GAL4/UAS system can be conditionally regulated using a temporal and regional gene expression targeting (TARGET) system that responds to heat shock induction. However heat shock-related temperature shifts sometimes cause unexpected physiological responses that confound behavioral analyses. We describe here the construction of a drug-inducible version of this system that takes advantage of tissue-specific GAL4 driver lines to yield either RU486-activated LexA-progesterone receptor chimeras (LexPR) or ß-estradiol-activated LexA-estrogen receptor chimeras (XVE). Upon induction, these chimeras bind to a LexA operator (LexAop) and activate transgene expression. Using GFP expression as a marker for induction in fly brain cells, both approaches are capable of tightly and precisely modulating transgene expression in a temporal and dosage-dependent manner. Additionally, tissue-specific GAL4 drivers resulted in target gene expression that was restricted to those specific tissues. Constitutive expression of the active PKA catalytic subunit using these systems altered the sleep pattern of flies, demonstrating that both systems can regulate transgene expression that precisely mimics regulation that was previously engineered using the GeneSwitch/UAS system. Unlike the limited number of GeneSwitch drivers, this approach allows for the usage of the multitudinous, tissue-specific GAL4 lines for studying temporal gene regulation and tissue-specific gene expression. Together, these new inducible systems provide additional, highly valuable tools available to study gene function in Drosophila.