Liposomes were generated by lipid film hydration as described (Kallert et al., 2015 (link)). Briefly, dimethyldioctadecylammonium (DDA; 0.3 mg/ml; Avanti Polar Lipids), D-(+)-trehalose 6,6′-dibehenate (TDB; 0.25 mg/ml; Avanti Polar Lipids, Sigma Aldrich) and L-α-phosphatidylcholine (PC; 0.9 mg/ml; Avanti Polar Lipids) were mixed in chloroform (VWR): methanol (Sigma-Aldrich; 9:1, v/v). The organic solvents were evaporated under nitrogen flow. Liposomes were formed by hydrating the lipidfilm in 500 μl 10 mM Tris-buffer (pH 7.4; Sigma-Aldrich) for 25 min at 57°C within between vortexing, as previously described (Kennerknecht et al., 2020 (link)). Angie1 or Angiogenin were added in a 1:1 mixture to the liposomes at a final concentration of 2.7 mM. The size of liposomes was determined by Nanoparticle Tracking Analysis (NTA) using a ZetaView TWIN (Particle Metrix, Inning, Germany). Samples were diluted in TRIS-buffer and videos of the light-refracting particles were recorded with the following settings: 25°C fixed temperature, 11 positions, 1 cycle, sensitivity 85, shutter 100, 15 fps, 2 s videos/position, and six measurements. The number and size distribution were evaluated by ZetaView Analyze (Version 08.05.05 SP2), as previously described (Conzelmann et al., 2020 (link)).
Zetaview twin
Manufactured by Particle Metrix
Sourced in Germany
The ZetaView TWIN is a laboratory instrument designed for the characterization of nanoparticles and colloids. It utilizes a combination of Dynamic Light Scattering (DLS) and Nanoparticle Tracking Analysis (NTA) techniques to measure the size, concentration, and zeta potential of samples. The instrument is capable of analyzing a wide range of materials, including proteins, polymers, liposomes, and other nanomaterials.
Automatically generated - may contain errors
Lab products found in correlation
Zetaview software, by Particle Metrix (3 mentions)
Chloroform, by Merck Group (1 mentions)
Methanol, by Merck Group (1 mentions)
L α phosphatidylcholine, by Avanti Polar Lipids (1 mentions)
Dimethyldioctadecylammonium, by Avanti Polar Lipids (1 mentions)
Pierce bca protein assay, by Thermo Fisher Scientific (1 mentions)
Zetaview, by Particle Metrix (1 mentions)
Zetaview software 8, by Particle Metrix (1 mentions)
13 protocols using zetaview twin
1
Liposome Preparation for Angiogenin Delivery
2
Isolation and Characterization of Extracellular Vesicles
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200,000 cells from the WT/FRα/FRα+CD9 lines adapted to SFM were seeded in 6-well plates (3 technical replicates per line). Cells were pelleted by centrifugation (1,000 rpm for 5 min) and the supernatant was further centrifuged at 2,000 g for 30 min to remove debris. Supernatant was then mixed with 0.5 volumes of Total Exosome Isolation reagent and samples were further processed as per manufacturer’s instructions. Final resuspension of EVs pellets was in 200 μl PBS. EVs concentration was measured with the ZetaView TWIN (Particle Metrix GmbH) using the 488 nm laser in scatter modality. Sensitivity was set to 80%, shutter to 100 units and temperature to 24°C. Samples were diluted 1:50 to 1:200 in PBS and recorded in triplicates (Measurement Mode: Size Distribution, 3 Cycles, 11 Positions). EVs parameters were calculated by the ZetaView software (Particle Metrix GmbH).
3
Nanoparticle Characterization by ZetaView
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Samples were diluted in 1 ml of ddH2O and concentration, size distribution, or surface charge (zeta potential) was measured 3 times using a ZetaView TWIN (Particle Metrix) with the software Zetaview (Zetaview Nanoparticle Tracking Analyzer RRID:SCR_016647). Zeta potential was calculated by electrophoretic mobility of the fibril samples.
4
Characterizing Skin-Derived Extracellular Vesicles
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Nanoparticle tracking analysis (NTA) was conducted with a Zetaview TWIN (Particle Metrix, Meerbusch, DE) to confirm the diameter and concentration of the extracellular vesicles. Lactobacillus species-derived extracellular vesicles (LpEVs) were isolated from human skin, suspended in filtered DW at 20.15 °C and were irradiated with a blue-light laser wavelength (λ = 488 nm). The sample conductivity was performed at 42.19 μS/cm and the filter wavelength was measured with backscatter detection. Samples were measured with dilution (dilution factor was 500) on the equivalent sample aliquot. The data were analyzed using ZetaView Software (version 8.05).
5
Nanoparticle Tracking Analysis of Plasma sEVs
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Nanoparticle tracking analysis (NTA) was performed on ZetaView® TWIN (Particle Metrix) to determine the size distribution and concentration of the isolated particles. Freshly isolated plasma sEVs from HNC patients were diluted up to 1:10,000 in PBS and from HD and NED up to 1:2000 in PBS and measured at room temperature (RT). Concentration and size ranges were calculated by ZetaView Software (Particle Metrix Version 8.05.11 SP1 and SP2, Sensitivity 80%, Shutter 100, 11 positions, 2 cycles).
6
Characterization of Extracellular Vesicles
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TEM of SEVs from HDs and HNC patients was performed at the Electron Microscopy Core Facility of Heidelberg University as previously described 20 .
SEVs were diluted up to 1:3000 in PBS and measured at room temperature (RT) using the Nanoparticle tracking analyzer (NTA) ZetaView® TWIN (Particle Metrix). Particle size and concentration were determined using the ZetaView Software (Particle Metrix Version 8.05.11 SP1 and SP2, Sensitivity 85%, Shutter 100, Min Area 15, Max Area 2000, Min Brightness 20, Trace Length 30, 11 positions, 2 cycles).
For protein quantification, Pierce™ BCA protein assay (Thermo Scientific) was performed according to the manufacturer’s instructions.
SEVs were diluted up to 1:3000 in PBS and measured at room temperature (RT) using the Nanoparticle tracking analyzer (NTA) ZetaView® TWIN (Particle Metrix). Particle size and concentration were determined using the ZetaView Software (Particle Metrix Version 8.05.11 SP1 and SP2, Sensitivity 85%, Shutter 100, Min Area 15, Max Area 2000, Min Brightness 20, Trace Length 30, 11 positions, 2 cycles).
For protein quantification, Pierce™ BCA protein assay (Thermo Scientific) was performed according to the manufacturer’s instructions.
7
NTA Analysis of Frozen Samples
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NTA was performed by the ZetaVIEW TWIN (Particle Metrix, Inning am Ammersee, Germany) using the software ZetaVIEW (Particle Metrix, version 8.05.14 SP7) and a temperature control to maintain constant 25 °C. The flow chamber was flushed with 10 mL PBS between each measurement. The samples were diluted in PBS in an appropriate ratio and measured at 11 positions with the following settings: camera sensitivity of 80 and shutter set to 100. Particle concentrations were calculated by multiplying with the used dilution factor. The samples were measured once on the day that they were collected. They were subsequently frozen, kept at −80 °C for 7 days, thawed, and then measured for a second time.
8
Characterizing Nanoparticle Corona Complexes
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The corona complexes
formed on 200 nm silica nanoparticle were characterized by nanoparticle
tracking analysis using a ZetaView TWIN instrument (Particle Metrix).
All samples were diluted in PBS to a final volume of 1 mL. Optimal
measurement concentrations were found by pretesting the particle per
frame value (100–200 particles/frame) and were typically a
dilution of 1:10 000 from 1 mg/mL corona complexes or nanoparticles
in PBS. For each sample, three technical replicates of one cycle were
performed by scanning 11 cell positions each and capturing 60 frames
per position (video setting: medium) under the following settings:
focus, autofocus; camera sensitivity, 75.0; shutter, 100; scattering
intensity, between 4 and 7; cell temperature, 25 °C. After capture,
the videos were analyzed by the built-in ZetaView Software 8.05.05
with specific analysis parameters, which are a maximum particle size
of 1000, a minimum particle size of 5, and a minimum particle brightness
of 50. Two independent measurements on separate corona–nanoparticle
complex preparations were performed.
formed on 200 nm silica nanoparticle were characterized by nanoparticle
tracking analysis using a ZetaView TWIN instrument (Particle Metrix).
All samples were diluted in PBS to a final volume of 1 mL. Optimal
measurement concentrations were found by pretesting the particle per
frame value (100–200 particles/frame) and were typically a
dilution of 1:10 000 from 1 mg/mL corona complexes or nanoparticles
in PBS. For each sample, three technical replicates of one cycle were
performed by scanning 11 cell positions each and capturing 60 frames
per position (video setting: medium) under the following settings:
focus, autofocus; camera sensitivity, 75.0; shutter, 100; scattering
intensity, between 4 and 7; cell temperature, 25 °C. After capture,
the videos were analyzed by the built-in ZetaView Software 8.05.05
with specific analysis parameters, which are a maximum particle size
of 1000, a minimum particle size of 5, and a minimum particle brightness
of 50. Two independent measurements on separate corona–nanoparticle
complex preparations were performed.
9
Nanoparticle Tracking Analysis of Extracellular Vesicles
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In order to confirm the morphology, including diameter and particle concentration, of the extracellular vesicles isolated by ultracentrifugation, nanoparticle tracking analysis was conducted with a ZetaView TWIN (Particle Metrix, Meerbusch, DE). PB-EVs (including L. holzapfelii isolated from the human scalp) suspended in filtered DPBS at 22.93 °C were irradiated with a blue-light laser wavelength (λ = 488 nm). The conductivity of samples was evaluated at 14.14 μS/cm, and filter wavelength was measured with backscatter detection. Samples were measured with dilution (dilution factor is 2000) and an average of 11 on the equivalent sample aliquot. Data were analyzed using ZetaView Software (version 8.05).
10
Characterizing Nanoparticles in Saliva and EV
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Nanoparticle tracking analysis (NTA) to determine the concentration and the size distribution of particles in saliva and EV isolations was performed using a ZetaView TWIN (Particle Metrix). Samples were diluted in particle‐free PBS and videos of the light‐refracting particles were recorded with the following settings: 25 °C fixed temperature, 11 positions, 1 cycle, sensitivity 85, shutter 100, 15 fps, 2 s videos/position, 3–5 measurements. The number, size distribution, and Zeta potential were evaluated by ZetaView Analyze 08.05.05 SP2. Between the samples, the chamber was thoroughly flushed with particle‐free PBS. To measure the apparent size of VLPs after incubation with compounds (60 min 37 °C), MLVgagYFP VLPs were tracked in 488 F‐NTA mode and sizes were determined by five acquisitions. For zeta potential analysis, VLPs incubated with compounds were diluted in ddH2O and zeta potential was measured in constant mode, five acquisitions in 488 nm F‐NTA per sample.
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