Highly Stable Plasmonic Nanostructures on a Nickel-Sputtered Glass and Polymeric Optical Fiber Sensors

Plasmonics - This study shows development of highly sensitive and stable localized surface plasmon resonance (LSPR)-active U-bent glass and polymeric optical fiber (GOF and POF) sensor probes by a...     

Automated SNP Genotype Clustering Algorithm to Improve Data Completeness in High-Throughput SNP Genotyping Datasets from Custom Arrays

High-throughput SNP genotyping platforms use automated genotype calling algo- rithms to assign genotypes. While these algorithms work efficiently for individual platforms, they are not compatible with other platforms, and have individual biases that result in missed genotype calls. Here we present data on the use of a second complementary SNP genotype clustering algorithm. The algorithm was originally designed for individual fluorescent SNP genotyping assays, and has been opti- mized to permit the clustering of large datasets generated from custom-designed Affymetrix SNP panels. In an analysis of data from a 3K array genotyped on 1,560 samples, the additional analysis increased the overall number of genotypes by over 45,000, significantly improving the completeness of the experimental data. This analysis suggests that the use of multiple genotype calling algorithms may be ad- visable in high-throughput SNP genotyping experiments. The software is written in Perl and is available from the corresponding author.     

A novel procedure for genotyping of single nucleotide polymorphisms in trisomy with genomic DNA and the invader assay

Individuals with trisomy 21 display complex phenotypes with differing degrees of severity. Numerous reliable methods have been established to diagnose the initial trisomy in these patients, but the identification and characterization of the genetic basis of the phenotypic variation in individuals with trisomy remains challenging. To date, methods that can accurately determine genotypes in trisomic DNA samples are expensive, require specialized equipment and complicated analyses. Here we report proof-of-concept results for an Invader® assay-based genotyping procedure that can determine SNP genotypes in trisomic genomic DNA samples in a simple and cost-effective manner. The procedure requires only two experimental steps: a real-time measurement of the fluorescent Invader® signal and analysis with a specifically designed clustering algorithm. The approach was tested using genomic DNA samples from 23 individuals with trisomy 21, and results were compared to genotypes previously determined with pyrosequencing. Additional assays for 15 SNPs were tested in a set of 21 DNA samples to assess assay performance. Our method successfully identified the correct SNP genotypes for the trisomic genomic DNA samples tested, and thus provides an alternative to determine SNP genotypes in trisomic DNA samples for subsequent association studies in patients with Down syndrome and other trisomies.     

Solution structure of copper ion-induced molecular aggregates of tyrosine melanin.

Melanin, the ubiquitous biological pigment, provides photoprotection by efficient filtration of light and also by its antioxidant behavior. In solutions of synthetic melanin, both optical and antioxidant behavior are affected by the aggregation states of melanin. We have utilized small-angle x-ray and neutron scattering to determine the molecular dimensions of synthetic tyrosine melanin in its unaggregated state in D(2)O and H(2)O to study the structure of melanin aggregates formed in the presence of copper ions at various copper-to-melanin molar ratios. In the absence of copper ions, or at low copper ion concentrations, tyrosine melanin is present in solution as a sheet-like particle with a mean thickness of 12.5 A and a lateral extent of approximately 54 A. At a copper-to-melanin molar ratio of 0.6, melanin aggregates to form long, rod-like structures with a radius of 32 A. At a higher copper ion concentration, with a copper-to-melanin ratio of 1.0, these rod-like structures further aggregate, forming sheet-like structures with a mean thickness of 51 A. A change in the charge of the ionizable groups induced by the addition of copper ions is proposed to account for part of the aggregation. The data also support a model for the copper-induced aggregation of melanin driven by pi stacking assisted by peripheral Cu(2+) complexation. The relationship between our results and a previous hypothesis for reduced cellular damage from bound-to-melanin redox metal ions is also discussed.     

Improving consensus structure by eliminating averaging artifacts

Background  

Common structural biology methods (i.e., NMR and molecular dynamics) often produce ensembles of molecular structures. Consequently, averaging of 3D coordinates of molecular structures (proteins and RNA) is a frequent approach to obtain a consensus structure that is representative of the ensemble. However, when the structures are averaged, artifacts can result in unrealistic local geometries, including unphysical bond lengths and angles.     

A microfluidic competitive immuno-aggregation assay for high sensitivity cell secretome detection

We report a high-sensitivity cell secretome detection method using competitive immuno-aggregation and a micro-Coulter counter. A target cell secretome protein competes with anti-biotin-coated microparticles (MPs) to bind with a biotinylated antibody (Ab), causing decreased aggregation of the functionalized MPs and formation of a mixture of MPs and aggregates. In comparison, without the target cell secretome protein, more microparticles are functionalized, and more aggregates are formed. Thus, a decrease in the average volume of functionalized microparticles/aggregates indicates an increase in cell secretome concentration. This volume change is measured by the micro-Coulter counter, which is used to quantitatively estimate the cell secretome concentration. Vascular endothelial growth factor (VEGF), one of the key cell secretome proteins that regulate angiogenesis and vascular permeabilization, was used as the target protein to demonstrate the sensing principle. A standard calibration curve was generated by testing samples with various VEGF concentrations. A detection range from 0.01 ng/mL to 100.00 ng/mL was achieved. We further demonstrated the quantification of VEGF concentration in exogenous samples collected from the secretome of human mesenchymal stem cells (hMSCs) at different incubation times. The results from the assay agree well with the results of a parallel enzyme-linked immunoabsorbent assay (ELISA) test, indicating the specificity and reliability of the competitive immuno-aggregation assay. With its simple structure and easy sample preparation, this assay not only enables high sensitivity detection of VEGF but also can be readily extended to other types of cell secretome analysis as long as the specific Ab is known.     

Comparison of linkage disequilibrium patterns between the HapMap CEPH samples and a family-based cohort of Northern European descent

The International HapMap Consortium has determined the linkage disequilibrium (LD) patterns of four major human populations. The aim of our investigation was to compare the LD patterns of the HapMap CEPH (Centre d'Etude du Polymorphisme Humain) samples with a family-based cohort of similar ancestry to determine its usefulness as a reference population for disease association studies. We examined four genomic regions on chromosomes 7q, 12p, and 14q totaling 14.3 Mb, initially identified in our linkage study of obesity and the metabolic syndrome. Near identical patterns of LD were detected in both populations. Furthermore, tagSNPs selected based on the HapMap CEPH cohort data capture over 98% of the variants at an r2 > 0.8 in the disease cohort. This confirms the usefulness of the CEPH cohort of the HapMap as a reference sample for further investigations into the genomic variation of populations of Northern European descent.