High throughput nanoparticle tracking analysis for monitoring outer membrane vesicle production.
TL;DR: This study shows that automated measurements in flow mode can be established with advanced scripting to reduce the workload for the user.
read more
Abstract: Outer membrane vesicles (OMVs) are spherical membrane nanoparticles released by Gram-negative bacteria. OMVs can be quantified in complex matrices by nanoparticle tracking analysis (NTA). NTA can be performed in static mode or with continuous sample flow that results in analysis of more particles in a smaller time-frame. Flow measurements must be performed manually despite the availability of a sample changer on the NanoSight system. Here we present a method for automated measurements in flow mode. OMV quantification in flow mode results in lower variance in particle quantification (coefficient of variation (CV) of 6%, CV static measurements of 14%). Sizing of OMVs was expected to be less favorable in flow mode due to the increased movement of the particles. However, we observed a CV of 3% in flow mode and a CV of 8% in static measurements. Flow rates of up to 5 µL/min displayed correct size and particle measurements, however, particle concentration was slightly lower than in static measurements. The automated method was used to assess OMV release of batch cultures of Neisseria meningitidis. The bacteria released more OMVs in stationary growth phase, while the size of the vesicles remained constant throughout the culture. Taken together, this study shows that automated measurements in flow mode can be established with advanced scripting to reduce the workload for the user.
read more
Chat with Paper
AI Agents for this Paper
Find similar papers on Google Scholar, PubMed and Arxiv
Write a critical review of this paper
Analyze citations of this paper to find unaddressed research gaps
Citations
Bacterial outer membrane vesicles as a platform for biomedical applications: An update.
TL;DR: The contributions of the structure and composition of OMV to their applications are analyzed, the methods used to isolate and characterize OMVs are summarized, and recent progress and future perspectives of OMVs in biomedical applications are highlighted.
231
Protocol for serum exosomal miRNAs analysis in prostate cancer patients treated with radiotherapy
TL;DR: The recovery of exosomal miRNAs and their differential expression after radiation treatment suggests promising biomarker potential that requires further investigation in larger patient cohorts.
Comparative Analysis of Outer Membrane Vesicle Isolation Methods With an Escherichia coli tolA Mutant Reveals a Hypervesiculating Phenotype With Outer-Inner Membrane Vesicle Content.
Shelby L. Reimer,Daniel R. Beniac,Shannon L. Hiebert,Timothy F. Booth,Patrick Chong,Garrett Westmacott,George G. Zhanel,Denice C. Bay +7 more
TL;DR: In this paper, two conventional techniques, ultracentrifugation (UC) and ultradiafiltration (UF), are used interchangeably to isolate OMVs, however, there is concern that each technique may inadvertently alter the properties of isolated OMVs during study, and to address this concern, they compared two OMV isolation methods, UC and UF, with respect to final OMV quantities, size distributions and morphologies using a hypervesiculating Escherichia coli K-12 ΔtolA mutant.
The Application of the CRISPR-Cas System in Antibiotic Resistance
01 Aug 2022
TL;DR: In this paper , the authors introduce the structure and working mechanism of CRISPR-Cas systems, followed by delivery strategies, and then focus on the relationship between antimicrobial resistance and CRISpl-Cas, and provide a reference for the prevention and control of the spread of antibiotic resistance.
51
Spontaneously released Neisseria meningitidis outer membrane vesicles as vaccine platform: production and purification
Matthias J.H. Gerritzen,Merijn L.M. Salverda,Dirk E. Martens,René H. Wijffels,René H. Wijffels,Michiel Stork +5 more
TL;DR: It is concluded that heterologous OMVs show potential as a vaccine platform by showing that production of well characterized OMVs containing heterology antigens is possible with high yields by combining high oxygen concentrations with an optimized purification process.
49
References
Critical Evaluation of Nanoparticle Tracking Analysis (NTA) by NanoSight for the Measurement of Nanoparticles and Protein Aggregates
TL;DR: NTA is a powerful characterization technique that complements DLS and is particularly valuable for analyzing polydisperse nanosized particles and protein aggregates.
Outer-membrane vesicles from Gram-negative bacteria: biogenesis and functions
TL;DR: This Review discusses recent advances in the study of OMVs, focusing on new insights into the mechanisms of biogenesis and the functions of these vesicles.
1.5K
Biological functions and biogenesis of secreted bacterial outer membrane vesicles.
Adam Kulp,Meta J. Kuehn +1 more
TL;DR: Comparisons of similar budding mechanisms from diverse biological domains have provided new insight into evaluating mechanisms for outer membrane vesiculation.
Sizing and phenotyping of cellular vesicles using Nanoparticle Tracking Analysis
R Dragovic,Chris Gardiner,Alexandra S. Brooks,Dionne Tannetta,David J. P. Ferguson,Patrick Hole,Bob Carr,Christopher W.G. Redman,Adrian L. Harris,Peter J. Dobson,Paul Harrison,Ian L. Sargent +11 more
TL;DR: By combining NTA with fluorescence measurement it is demonstrated that vesicles can be labeled with specific antibody-conjugated quantum dots, allowing their phenotype to be determined, demonstrating that NTA is far more sensitive than conventional flow cytometry.
1.3K
Particle size distribution of exosomes and microvesicles determined by transmission electron microscopy, flow cytometry, nanoparticle tracking analysis, and resistive pulse sensing.
E. van der Pol,Frank A. W. Coumans,Anita E. Grootemaat,Chris Gardiner,I.L. Sargent,Paul Harrison,A. Sturk,T. G. van Leeuwen,Rienk Nieuwland +8 more
TL;DR: Enumeration of extracellular vesicles has clinical potential as a biomarker for disease and currently employed techniques detect concentrations ranging from 104 to 1012 vesicle mL–1.
854