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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV210210	1/1	Homo sapiens	Expi293F	(d)(U)C Tangential flow filtration qEV10 Filtration UF	Driedonks T	2022	63%
Study summary Full title Pharmacokinetics and biodistribution of extracellular vesicles administered intravenously and intranasally to . All authors Driedonks T, Jiang L, Carlson B, Han Z, Liu G, Queen SE, Shirk EN, Gololobova O, Liao Z, Nyberg LH, Lima G, Paniushkina L, Garcia-Contreras M, Schonvisky K, Castell N, Stover M, Guerrero-Martin S, Richardson R, Smith B, Machairaki V, Lai CP, Izzi JM, Hutchinson EK, Pate KAM, Witwer KW Journal J Extracell Biol Abstract Extracellular vesicles (EVs) have potential in disease treatment since they can be loaded with thera (show more...)Extracellular vesicles (EVs) have potential in disease treatment since they can be loaded with therapeutic molecules and engineered for retention by specific tissues. However, questions remain on optimal dosing, administration, and pharmacokinetics. Previous studies have addressed biodistribution and pharmacokinetics in rodents, but little evidence is available for larger animals. Here, we investigated the pharmacokinetics and biodistribution of Expi293F-derived EVs labelled with a highly sensitive nanoluciferase reporter (palmGRET) in a non-human primate model (Macaca nemestrina), comparing intravenous (IV) and intranasal (IN) administration over a 125-fold dose range. We report that EVs administered IV had longer circulation times in plasma than previously reported in mice and were detectable in cerebrospinal fluid (CSF) after 30-60 minutes. EV association with PBMCs, especially B-cells, was observed as early as one minute post-administration. EVs were detected in liver and spleen within one hour of IV administration. However, IN delivery was minimal, suggesting that pretreatment approaches may be needed in large animals. Furthermore, EV circulation times strongly decreased after repeated IV administration, possibly due to immune responses and with clear implications for xenogeneic EV-based therapeutics. We hope that our findings from this baseline study in macaques will help to inform future research and therapeutic development of EVs. (hide) EV-METRIC 63% (92nd percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin pLenti-palmGRET (Addgene #158221) transient expression Focus vesicles extracellular vesicle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (Differential) (ultra)centrifugation Tangential flow filtration Commercial method Filtration Ultrafiltration Protein markers EV: CD9/ CD63/ CD81/ TSG101/ CD29/ CD146/ CD326/ CD44/ ROR1/ CD24 non-EV: Calnexin Proteomics no Show all info Study aim Biodistribution/pharmacokinetics/Mechanism of uptake/transfer Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Expi293F Cell count 3.30E+09 Separation Method Filtration steps 0.2 or 0.22 µm Ultra filtration Cut-off size (kDa) 100 Membrane type Regenerated cellulose Commercial kit qEV10 Other Name other separation method Tangential flow filtration Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins CD9/ CD63/ TSG101/ CD47 Not detected contaminants Calnexin Other 1 SP-IRIS (Exoview) Detected EV-associated proteins CD63/ CD81 Other 2 MACSPlex Detected EV-associated proteins CD9/ CD63/ CD81/ CD29/ CD146/ CD326/ CD44/ ROR1/ CD24 Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Mean Reported size (nm) 122.6 EV concentration Yes Particle analysis: flow cytometry Flow cytometer type Amnis ImageStream MKII Hardware adjustment EM EM-type Transmission-EM Image type Wide-field Extra information Size-exclusion chromatography details: Izon qEV10, first 20 ml after the void fraction were considered EV-enriched fractions/ Tangential Flow Filtration: Sartorius Vivaflow 50R cassettes, 100 kD

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EV-TRACK ID	EV210210
species	Homo sapiens
sample type	Cell culture
cell type	Expi293F
condition	pLenti-palmGRET (Addgene #158221) transient expression
separation protocol	dUC/ Tangential flow filtration/ qEV10/ Filtration/ Ultrafiltration
Exp. nr.	1
EV-METRIC %	63