Asymmetrical flow field-flow fractionation technique for separation and characterization of biopolymers and bioparticles

Yohannes, Gebrenegus; Jussila, Matti; Hartonen, Kari; Riekkola, Marja‐Liisa

doi:10.1016/j.chroma.2010.12.110

Cited by 136 publications

(73 citation statements)

References 106 publications

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“…AFFF exploits the parabolic flow profile created by the laminar flow of a sample through a thin, parallel plate flow channel, where the lower surface is solvent permeable [122]. A perpendicular force applied to the laminar flow stream drives molecules towards the permeable boundary layer of the channel [123].…”

Section: Additional Techniques Utilized For Aβ Aggregate Size Detementioning

confidence: 99%

“…A perpendicular force applied to the laminar flow stream drives molecules towards the permeable boundary layer of the channel [123]. Because Brownian motion of the particles creates a counteracting force, smaller particles localize higher in the channel leading to separation of different molecular sizes, with smaller molecules eluting first [122]. Rambaldi et al utilized AFFF-MALS to monitor the aggregation of Aβ 1-42 in PBS (pH 7.4) at room temperature over 24 h [119].…”

Section: Additional Techniques Utilized For Aβ Aggregate Size Detementioning

confidence: 99%

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Unraveling the Early Events of Amyloid-β Protein (Aβ) Aggregation: Techniques for the Determination of Aβ Aggregate Size

Pryor

Moss

Hestekin

2012

IJMS

102

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The aggregation of proteins into insoluble amyloid fibrils coincides with the onset of numerous diseases. An array of techniques is available to study the different stages of the amyloid aggregation process. Recently, emphasis has been placed upon the analysis of oligomeric amyloid species, which have been hypothesized to play a key role in disease progression. This paper reviews techniques utilized to study aggregation of the amyloid-β protein (Aβ) associated with Alzheimer’s disease. In particular, the review focuses on techniques that provide information about the size or quantity of oligomeric Aβ species formed during the early stages of aggregation, including native-PAGE, SDS-PAGE, Western blotting, capillary electrophoresis, mass spectrometry, fluorescence correlation spectroscopy, light scattering, size exclusion chromatography, centrifugation, enzyme-linked immunosorbent assay, and dot blotting.

show abstract

Section: Additional Techniques Utilized For Aβ Aggregate Size Detementioning

confidence: 99%

Section: Additional Techniques Utilized For Aβ Aggregate Size Detementioning

confidence: 99%

Unraveling the Early Events of Amyloid-β Protein (Aβ) Aggregation: Techniques for the Determination of Aβ Aggregate Size

Pryor

Moss

Hestekin

2012

IJMS

102

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“…But, first the combination of flow field-flow fractionation with on-line multi-angle light scattering (A4F-MALLS; Wyatt, 1998) enabled scientists to gain a fast, detailed, and reliable quantitative insight into size distributions of submicron particles. It took another decade to establish A4F-MALLS for liposome size-analysis, providing now a level of resolution and precision not seen before with routine methods (Arifin & Palmer, 2003;Hong et al, 2010;Hupfeld et al, 2006Hupfeld et al, , 2009aHupfeld et al, ,b, 2010Jahn et al, 2007;Kuntsche et al, 2010Kuntsche et al, , 2012Yohannes et al, 2011).…”

Section: Introductionmentioning

confidence: 97%

Filter-extruded liposomes revisited: a study into size distributions and morphologies in relation to lipid-composition and process parameters

Hinna

Steiniger

Hupfeld³

et al. 2015

Journal of Liposome Research

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Filter-extrusion is a widely used technique for down-sizing of phospholipid vesicles. In order to gain a detailed insight into size and size distributions of filter-extruded vesicles composed of egg phosphatidyl-choline (with varying fractions of cholesterol)--in relation to extrusion-parameters (pore-size, number of filter passages, and flow-rate), flow field-flow fractionation in conjunction with multi-angle laser light scattering (AF4-MALLS, Wyatt Technology Corp., Santa Barbara, CA) was employed. Liposome size-distributions determined by AF4-MALLS were compared with those of dynamic light scattering and correlated with cryo-transmission electron microscopy and (31)P-NMR-analysis of lamellarity. Both the mean size of liposome and the width of size distribution were found to decrease with sequential extrusion through smaller pore size filters, starting at a size range of ≈70-415 nm upon repeated extrusion through 400 nm pore-filters, eventually ending with a size range from ≈30 to 85 nm upon extrusion through 30 nm pore size filters. While for small pores sizes (50 nm), increased flow rates resulted in smaller vesicles, no significant influence of flow rate on mean vesicle size was seen with larger pores. Cholesterol at increasing mol fractions up to 0.45 yielded bigger vesicles (at identical process conditions). For a cholesterol mol fraction of 0.5 in combination with small filter pore size, a bimodal size distribution was seen indicating cholesterol micro-crystallites. Finally, a protocol is suggested to prepare large (∼ 300 nm) liposomes with rather narrow size distribution, based on the filter extrusion at defined flow-rates in combination with freeze-/thaw-cycling and bench-top centrifugation.

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“…The state-of-the-art technology, asymmetric-flow field-flow fractionation (AF4) 8 , exhibits unique capability to separate nanoparticles and has been widely utilized to characterize nanoparticles and polymers in the pharmaceutical industry and to examine various biological macromolecules, protein complexes and viruses 8, 9 , but rarely tested for extracellular vesicle (EV) analysis 10-14 . Using AF4, nanoparticles are separated based on their density and hydrodynamic properties by two perpendicular flows, i.e., the forward laminar channel flow and the variable crossflow.…”

Section: Introductionmentioning

confidence: 99%

Identification of distinct nanoparticles and subsets of extracellular vesicles by asymmetric flow field-flow fractionation

et al. 2018

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The heterogeneity of exosomal populations has hindered our understanding of their biogenesis, molecular composition, biodistribution, and functions. By employing asymmetric-flow field-flow fractionation (AF4), we identified two exosome subpopulations (large exosome vesicles, Exo-L, 90-120 nm; small exosome vesicles, Exo-S, 60-80 nm) and discovered an abundant population of non-membranous nanoparticles termed “exomeres” (~35 nm). Exomere proteomic profiling revealed an enrichment in metabolic enzymes and hypoxia, microtubule and coagulation proteins and specific pathways, such as glycolysis and mTOR signaling. Exo-S and Exo-L contained proteins involved in endosomal function and secretion pathways, and mitotic spindle and IL-2/STAT5 signaling pathways, respectively. Exo-S, Exo-L, and exomeres each had unique N-glycosylation, protein, lipid, and DNA and RNA profiles and biophysical properties. These three nanoparticle subsets demonstrated diverse organ biodistribution patterns, suggesting distinct biological functions. This study demonstrates that AF4 can serve as an improved analytical tool for isolating and addressing the complexities of heterogeneous nanoparticle subpopulations.

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Asymmetrical flow field-flow fractionation technique for separation and characterization of biopolymers and bioparticles

Cited by 136 publications

References 106 publications

Unraveling the Early Events of Amyloid-β Protein (Aβ) Aggregation: Techniques for the Determination of Aβ Aggregate Size

Unraveling the Early Events of Amyloid-β Protein (Aβ) Aggregation: Techniques for the Determination of Aβ Aggregate Size

Filter-extruded liposomes revisited: a study into size distributions and morphologies in relation to lipid-composition and process parameters

Identification of distinct nanoparticles and subsets of extracellular vesicles by asymmetric flow field-flow fractionation

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