Channel Blocking of MspA Revisited

Perera, Ayomi S.; Wang, Hongwang; Basel, Matthew T.; Pokhrel, Megh Raj; Gamage, Pubudu; Kalita, Mausam; Wendel, Sebastian O.; Sears, B.; Welideniya, Dhanushi; Liu, Yao; Turró, Claudia; Troyer, Deryl L.; Bossmann, Stefan H.

doi:10.1021/la3037296

Cited by 10 publications

(11 citation statements)

References 27 publications

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“…The zeta potentials of BAL exosomes were measured using a ZetaPALS Zeta Potential Analyzer (Brookhaven Instruments Corporation, Holtsville, NY, USA) and previously established procedures. [36] It was not necessary to adjust the exosome concentration or the pH of the solution. The first set of exosomes was measured directly at 18°C.…”

Section: Methodsmentioning

confidence: 99%

Effects of storage temperature on airway exosome integrity for diagnostic and functional analyses

Maroto

Zhao

Jamaluddin

et al. 2017

J of Extracellular Vesicle

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225

165

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Background: Extracellular vesicles contain biological molecules specified by cell-type of origin and modified by microenvironmental changes. To conduct reproducible studies on exosome content and function, storage conditions need to have minimal impact on airway exosome integrity.Aim: We compared surface properties and protein content of airway exosomes that had been freshly isolated vs. those that had been treated with cold storage or freezing.Methods: Mouse bronchoalveolar lavage fluid (BALF) exosomes purified by differential ultracentrifugation were analysed immediately or stored at +4°C or −80°C. Exosomal structure was assessed by dynamic light scattering (DLS), transmission electron microscopy (TEM) and charge density (zeta potential, ζ). Exosomal protein content, including leaking/dissociating proteins, were identified by label-free LC-MS/MS.Results: Freshly isolated BALF exosomes exhibited a mean diameter of 95 nm and characteristic morphology. Storage had significant impact on BALF exosome size and content. Compared to fresh, exosomes stored at +4°C had a 10% increase in diameter, redistribution to polydisperse aggregates and reduced ζ. Storage at −80°C produced an even greater effect, resulting in a 25% increase in diameter, significantly reducing the ζ, resulting in multilamellar structure formation. In fresh exosomes, we identified 1140 high-confidence proteins enriched in 19 genome ontology biological processes. After storage at room temperature, 848 proteins were identified. In preparations stored at +4°C, 224 proteins appeared in the supernatant fraction compared to the wash fractions from freshly prepared exosomes; these proteins represent exosome leakage or dissociation of loosely bound “peri-exosomal” proteins. In preparations stored at −80°C, 194 proteins appeared in the supernatant fraction, suggesting that distinct protein groups leak from exosomes at different storage temperatures.Conclusions: Storage destabilizes the surface characteristics, morphological features and protein content of BALF exosomes. For preservation of the exosome protein content and representative functional analysis, airway exosomes should be analysed immediately after isolation.

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Section: Methodsmentioning

confidence: 99%

Effects of storage temperature on airway exosome integrity for diagnostic and functional analyses

Maroto

Zhao

Jamaluddin

et al. 2017

J of Extracellular Vesicle

Self Cite

225

165

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“…Zeta potential measurements were performed using the ZetaPALS Zeta Potential Analyzer (Brookhaven Instruments Corporation), as described previously [ 24 ]. M .…”

Section: Methodsmentioning

confidence: 99%

Cell Based Drug Delivery: Micrococcus luteus Loaded Neutrophils as Chlorhexidine Delivery Vehicles in a Mouse Model of Liver Abscesses in Cattle

et al. 2015

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The recent WHO report on antibiotic resistances shows a dramatic increase of microbial resistance against antibiotics. With only a few new antibiotics in the pipeline, a different drug delivery approach is urgently needed. We have obtained evidence demonstrating the effectiveness of a cell based drug delivery system that utilizes the innate immune system as targeting carrier for antibacterial drugs. In this study we show the efficient loading of neutrophil granulocytes with chlorhexidine and the complete killing of E. coli as well as Fusobacterium necrophorum in in-vitro studies. Fusobacterium necrophorum causes hepatic abscesses in cattle fed high grain diets. We also show in a mouse model that this delivery system targets infections of F. necrophorum in the liver and reduces the bacterial burden by an order of magnitude from approximately 2•106 to 1•105.

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“…Zeta potential measurements were performed using ZetaPALS Zeta Potential Analyzer purchased from Brookhaven Instruments Corporation. 18 The HMNPs were characterized in 1Â PBS at 298 K. Their concentration was 0.1 mg mL À1 .…”

Section: Zeta Potential Measurementsmentioning

confidence: 99%

Hexagonal magnetite nanoprisms: preparation, characterization and cellular uptake

et al. 2015

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Channel Blocking of MspA Revisited

Cited by 10 publications

References 27 publications

Effects of storage temperature on airway exosome integrity for diagnostic and functional analyses

Effects of storage temperature on airway exosome integrity for diagnostic and functional analyses

Cell Based Drug Delivery: Micrococcus luteus Loaded Neutrophils as Chlorhexidine Delivery Vehicles in a Mouse Model of Liver Abscesses in Cattle

Hexagonal magnetite nanoprisms: preparation, characterization and cellular uptake

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