Biology subjects

Mucci, A.

Publications and source records attributed to Mucci, A..

3 recordsLinked to original sources

bioRxiv2025

Quantification of horizontal and vertical distribution of junctional proteins in fixed epithelial cells

Polarized epithelial cells form a tightly packed monolayer where individual cells are connected by cell-cell junctions, including tight junctions (TJ) and adherens junctions (AJ). Here, we present techniques for quantifying the horizontal and vertical distribution of junctional proteins in confluent, fixed epithelial cells. This approach is utilized to evaluate variations in the intensity and localization of the proteins that compose the AJ and TJ under different experimental conditions. Although our protocol is optimized for Madin-Darby Canine Kidney (MDCK) cells, it is adaptable to any cell line capable of forming cell-cell junctions. For complete details on the use and execution of this protocol, please refer to Rabino et al. (2024). 1 Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=190 HEIGHT=200 SRC="FIGDIR/small/651687v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@1d1db4forg.highwire.dtl.DTLVardef@954ea1org.highwire.dtl.DTLVardef@5d43eborg.highwire.dtl.DTLVardef@1162077_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2024

Scalable assessment of genome editing off-targets associated with genetic variants

Genome editing with RNA-guided DNA binding factors carries risk of off-target editing at homologous sequences. Genetic variants may introduce sequence changes that increase homology to a genome editing target, thereby increasing risk of off-target editing. Conventional methods to verify candidate off-targets rely on access to cells with genomic DNA carrying these sequences. However, for candidate off-targets associated with genetic variants, appropriate cells for experimental verification may not be available. Here we develop a method, Assessment By Stand-in Off-target LentiViral Ensemble with sequencing (ABSOLVE-seq), to integrate a set of candidate off-target sequences along with unique molecular identifiers (UMIs) in genomes of primary cells followed by clinically relevant gene editor delivery. Gene editing of dozens of candidate off-target sequences may be evaluated in a single experiment with high sensitivity, precision, and power. We provide an open-source pipeline to analyze sequencing data. This approach enables experimental assessment of the influence of human genetic diversity on specificity evaluation during gene editing therapy development.

genomics↗

bioRxiv2022

Epitope Mapping of SARS-CoV-2 Spike Protein Reveals Distinguishable Antibody Binding Activity of Vaccinated and Infected Individuals.

Previous studies have attempted to characterize the antibody response of individuals to the SARS-CoV-2 virus on a linear peptide level by utilizing peptide microarrays. These studies have helped to identify epitopes that have potential to be used for diagnostic tests to identify infected individuals, however, the immunological responses of individuals who have received the currently available Moderna mRNA-1273 or Pfizer BNT162b2 mRNA vaccines have not been characterized. We aimed to identify linear peptides of the SARS-CoV-2 spike protein that elicited high IgG or IgA binding activity and to compare the immunoreactivity of infected individuals to those who received both doses of either vaccines by utilizing peptide microarrays. Our results revealed peptide epitopes of significant IgG binding among recently infected individuals. Some of these peptides are located near functional domains implicated in the high infectivity of SARS-CoV-2. Vaccinated individuals lacked these distinct markers despite overall binding activity being similar.

immunology↗