Shinichi Utsunomiya

Large-scale microfabricated channel plates for high-throughput, fully automated DNA sequencing.

We have described a new DNA sequencing platform based on the Sanger chemistry, in which the large‐scale microfabricated channel plates and electrophoretic system result in higher‐throughput DNA sequencing. Three hundred and eighty‐four channels are arranged in a fan‐like shape on a 25×47 cm glass plate, on which 384 oval sample holes are connected to each channel coupled to the opposite anode access holes. Two microfabricated plates are set on the sequencing apparatus, in which sequencing electrophoresis is conducted on one plate and the preparation process is on another plate. Each sample hole is loaded with 2.3 μL volume of sample and injected into separation channels electrokinetically. High‐quality sequencing data were acquired using the pUC18 template, achieving an average read‐length of 1001 bases with 99% accuracy and a throughput of 5 Mbases per day per instrument. To assess the performance in actual sequencing field, the bacterial artificial chromosome shotgun library of the Pseudorca crassidens genome was sequenced, using 1/80 of the quantity of Sanger reagent (0.1 μL). We believe that this is the first demonstration of the useful performance of DNA sequencing using monolithic microfabricated devices with walk‐away operation.

A simple peak detection and label-free quantitation algorithm for chromatography-mass spectrometry

BackgroundLabel-free quantitation of mass spectrometric data is one of the simplest and least expensive methods for differential expression profiling of proteins and metabolites. The need for high accuracy and performance computational label-free quantitation methods is still high in the biomarker and drug discovery research field. However, recent most advanced types of LC-MS generate huge amounts of analytical data with high scan speed, high accuracy and resolution, which is often impossible to interpret manually. Moreover, there are still issues to be improved for recent label-free methods, such as how to reduce false positive/negatives of the candidate peaks, how to expand scalability and how to enhance and automate data processing. AB3D (A simple label-free quantitation algorithm for Biomarker Discovery in Diagnostics and Drug discovery using LC-MS) has addressed these issues and has the capability to perform label-free quantitation using MS1 for proteomics study.ResultsWe developed an algorithm called AB3D, a label free peak detection and quantitative algorithm using MS1 spectral data. To test our algorithm, practical applications of AB3D for LC-MS data sets were evaluated using 3 datasets. Comparisons were then carried out between widely used software tools such as MZmine 2, MSight, SuperHirn, OpenMS and our algorithm AB3D, using the same LC-MS datasets. All quantitative results were confirmed manually, and we found that AB3D could properly identify and quantify known peptides with fewer false positives and false negatives compared to four other existing software tools using either the standard peptide mixture or the real complex biological samples of Bartonella quintana (strain JK31). Moreover, AB3D showed the best reliability by comparing the variability between two technical replicates using a complex peptide mixture of HeLa and BSA samples. For performance, the AB3D algorithm is about 1.2 - 15 times faster than the four other existing software tools.ConclusionsAB3D is a simple and fast algorithm for label-free quantitation using MS1 mass spectrometry data for large scale LC-MS data analysis with higher true positive and reasonable false positive rates. Furthermore, AB3D demonstrated the best reproducibility and is about 1.2- 15 times faster than those of existing 4 software tools.

Automatic left ventricular volume measurements on contrast-enhanced ultrafast cine magnetic resonance imaging

To assess the accuracy of automatic extraction of the left ventricular inner contour on contrast-enhanced ultrafast cine magnetic resonance (MR) images, we compared the values obtained by this method with those obtained using intravenous digital subtraction left ventriculography. High-quality single breath-hold contrast-enhanced ultrafast cine MR images were obtained in all cardiac phases on horizontal and vertical long axis sections of the left ventricle. For ultrafast cine MR imaging, a phase-rewind gradient-echo (rewind-SMASH) sequence was used. Automatic extraction of the left ventricular inner contour on contrast-enhanced ultrafast cine MR images was performed in all cardiac phases. High-quality left ventricular images of the horizontal long axis section were obtained in 127 of 160 patients (79%). The automatic extraction of the left ventricular contour was easily performed on high-quality images with very short processing time (4 s/frame). The values for left ventricular volumes obtained with the automatic extraction method on contrast-enhanced ultrafast cine MR imaging were correlated well with those obtained with the manual extraction method and IV-DSA in high quality cardiac images. The biplane modified Simpsons method using automatic extraction is an accurate and highly reproducible method for evaluating left ventricular volumes.

Peptide Peak Detection for Low Resolution MALDI-TOF Mass Spectrometry

A new peak detection method has been developed for rapid selection of peptide and its fragment ion peaks for protein identification using tandem mass spectrometry. The algorithm applies classification of peak intensities present in the defined mass range to determine the noise level. A threshold is then given to select ion peaks according to the determined noise level in each mass range. This algorithm was initially designed for the peak detection of low resolution peptide mass spectra, such as matrix-assisted laser desorption/ionization Time-of-Flight (MALDI-TOF) mass spectra. But it can also be applied to other type of mass spectra. This method has demonstrated obtaining a good rate of number of real ions to noises for even poorly fragmented peptide spectra. The effect of using peak lists generated from this method produces improved protein scores in database search results. The reliability of the protein identifications is increased by finding more peptide identifications. This software tool is freely available at the Mass++ home page (http://www.first-ms3d.jp/english/achievement/software/).