Biology subjects

Wu, V.

Publications and source records attributed to Wu, V..

3 recordsLinked to original sources

bioRxiv2021

Mass recalibration for desorption electrospray ionization mass spectrometry imaging using endogenous reference ions.

1BackgroundMass spectrometry imaging (MSI) data often consist of tens of thousands of mass spectra collected from a sample surface. During the time necessary to perform a single acquisition, it is likely that uncontrollable factors alter the validity of the initial mass calibration of the instrument, resulting in mass errors of magnitude significantly larger than their expected values. This phenomenon has a two-fold detrimental effect: a) it reduces the ability to interpret the results based on the observed signals, b) it can affect the quality of the observed signal spatial distributions. ResultsWe present a post-acquisition computational method capable of reducing the observed mass drift by up to 60 ppm in biological samples, exploiting the presence of typical molecules with a known mass-to-charge ratio. The procedure, tested on time-of-flight (TOF) and Orbitrap mass spectrometry analyzers interfaced to a desorption electrospray ionization (DESI) source, improves the molecular annotation quality and the spatial distributions of the detected ions. ConclusionThe presented method represents a robust and accurate tool for performing post-acquisition mass recalibration of DESI-MSI datasets and can help to increase the reliability of the molecular assignment and the data quality.

bioinformatics↗

bioRxiv2021

Genome-Wide Identification of 5-Methylcytosine Sites in Bacterial Genomes By High-Throughput Sequencing of MspJI Restriction Fragments

Single-molecule Real-Time (SMRT) sequencing can easily identify sites of N6-methyladenine and N4-methylcytosine within DNA sequences, but similar identification of 5-methylcytosine sites is not as straightforward. In prokaryotic DNA, methylation typically occurs within specific sequence contexts, or motifs, that are a property of the methyltransferases that "write" these epigenetic marks. We present here a straightforward, cost-effective alternative to both SMRT and bisulfite sequencing for the determination of prokaryotic 5-methylcytosine methylation motifs. The method, called MFRE-Seq, relies on excision and isolation of fully methylated fragments of predictable size using MspJI-Family Restriction Enzymes (MFREs), which depend on the presence of 5-methylcytosine for cleavage. We demonstrate that MFRE-Seq is compatible with both Illumina and Ion Torrent sequencing platforms and requires only a digestion step and simple column purification of size-selected digest fragments prior to standard library preparation procedures. We applied MFRE-Seq to numerous bacterial and archaeal genomic DNA preparations and successfully confirmed known motifs and identified novel ones. This method should be a useful complement to existing methodologies for studying prokaryotic methylomes and characterizing the contributing methyltransferases.

genomics↗

bioRxiv2020

The identity of human tissue-emigrant CD8+ T cells

Lymphocyte migration is essential for human adaptive immune surveillance. However, our current understanding of this process is rudimentary, because most human studies to date have been restricted to immunological analyses of blood and various tissues. To address this issue, we used an integrated approach to characterize tissue-emigrant immune cells in thoracic duct lymph (TDL). In humans and non-human primates, lymphocytes were by far the most abundant immune lineage population in efferent lymph, and a vast majority of these lymphocytes were T cells. Cytolytic CD8+ T cell subsets were clonotypically discrete and selectively confined to the intravascular circulation, persisting for months after inhibition of S1P-dependent tissue egress by FTY-720. In contrast, non-cytolytic CD8+ T cell subsets with stem-like epigenetic and transcriptional signatures predominated in tissues and TDL. Collectively, these data provide an atlas of the migratory immune system and define the nature of tissue-emigrant CD8+ T cells that recirculate via TDL.

immunology↗