scholarly journals Elucidating Methods for Isolation and Quantification of Exosomes: A Review

2021 ◽  
Author(s):  
Talitha Keren Kurian ◽  
Soumyabrata Banik ◽  
Dharshini Gopal ◽  
Shweta Chakrabarti ◽  
Nirmal Mazumder
Keyword(s):  
Gold Standard ◽  
Tumor Metastasis ◽  
Biological Fluids ◽  
Size Exclusion ◽  
Nanoparticle Tracking Analysis ◽  
Standard Methods ◽  
Advantages And Disadvantages ◽  
Resistive Pulse ◽  
Tunable Resistive Pulse Sensing ◽  
Exclusion Chromatography

AbstractExosomes are the smallest extracellular vesicles present in most of the biological fluids. They are found to play an important role in cell signaling, immune response, tumor metastasis, etc. Studies have shown that these vesicles also have diagnostic and therapeutic roles for which their accurate detection and quantification is essential. Due to the complexity in size and structure of exosomes, even the gold standard methods face challenges. This comprehensive review discusses the various standard methods such as ultracentrifugation, ultrafiltration, size-exclusion chromatography, precipitation, immunoaffinity, and microfluidic technologies for the isolation of exosomes. The principle of isolation of each method is described, as well as their specific advantages and disadvantages. Quantification of exosomes by nanoparticle tracking analysis, flow cytometry, tunable resistive pulse sensing, electron microscopy, dynamic light scattering, and microfluidic devices are also described, along with the applications of exosomes in various biomedical domains.

scholarly journals MAGNETIC NANOLIPOSOMES: A REVIEW ON PREPARATION AND CHARACTERIZATION METHODS

2021 ◽  
Vol 11 (JAN-APR/MAY-AUG/SEP-DEC) ◽  
pp. 1-8
Author(s):  
Palanisamy Selvamani ◽  
◽  
Venkata Krishnan Par ◽  
Rosi Rajamanickam ◽  
Subbiah Latha ◽  
...  
Keyword(s):  
Targeted Drug Delivery ◽  
Size Exclusion ◽  
Pharmacokinetic Parameters ◽  
Nanoparticle Tracking Analysis ◽  
Chemical Methods ◽  
Characterization Methods ◽  
Exclusion Chromatography ◽  
Characterization Studies ◽  
Targeted Drug ◽  
Co Precipitation

Magnetic nanoliposomes are versatile nanocarriers for targeted drug delivery. These liposomes can enhance the efficacy of bioactive compounds by improving pharmacokinetic parameters. Chemical ingredients of magnetic nanoliposomes include ferrofluid, cholesterol and phospholipid molecules. The preparation of magnetic nanoliposomes involves two steps viz. synthesis of magnetic nanoparticles and then combination with prepared nanoliposomes. The magnetic nanoliposomes are synthesized by various chemical methods including co-precipitation, thermal decomposition, microemulsion and hydrothermal synthesis. The nanoliposomes are prepared by sonication, extrusion, micro-fluidization techniques. Characterization studies included size, nanoparticle tracking analysis, flow cytometry, size exclusion chromatography, DSC, encapsulation efficiency. This study provides an overview of the preparation and characterization techniques of magnetic nanoliposomes.

scholarly journals Asymmetric depth-filtration - a versatile and scalable approach for isolation and purification of extracellular vesicles

2021 ◽  
Author(s):  
Vasiliy S. Chernyshev ◽  
Roman N. Chuprov-Netochin ◽  
Ekaterina Tsydenzhapova ◽  
Elena Victorovna Svirshchevskaya ◽  
Rimma A. Poltavtseva ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Biological Samples ◽  
Biological Fluids ◽  
Scale Up ◽  
Size Exclusion ◽  
Standard Laboratory ◽  
Laboratory Equipment ◽  
Isolation And Purification ◽  
Depth Filtration ◽  
Exclusion Chromatography

A novel asymmetric depth filtration (DF) approach for isolation of extracellular vesicles (EVs) from biological fluids is presented, and its performance is compared with established methods. The developed workflow is simple, inexpensive, and relatively fast. Compared with ultracentrifugation and size-exclusion chromatography, the developed method isolates EVs with higher purity and yield. Only standard laboratory equipment is needed for its implementation, which makes it suitable for low-resource locations. The described implementation of the method is suitable for EV isolation from small biological samples in diagnostic and treatment guidance applications. Following the scale-up routes adopted in the biomanufacturing of therapeutics, which routinely rely on DF as one of the product purification steps, the developed method may be scaled up to harvesting therapeutic EVs from large volumes of growth medium.

scholarly journals A Review of Exosomal Isolation Methods: Is Size Exclusion Chromatography the Best Option?

2020 ◽  
Vol 21 (18) ◽  
pp. 6466 ◽  
Author(s):  
Karim Sidhom ◽  
Patience O. Obi ◽  
Ayesha Saleem
Keyword(s):  
Size Exclusion Chromatography ◽  
Culture Media ◽  
Biological Fluids ◽  
Vital Role ◽  
Size Exclusion ◽  
Isolation Techniques ◽  
Exclusion Chromatography ◽  
Specialized Equipment ◽  
Pros And Cons ◽  
Poly Ethylene

Extracellular vesicles (EVs) are membranous vesicles secreted by both prokaryotic and eukaryotic cells and play a vital role in intercellular communication. EVs are classified into several subtypes based on their origin, physical characteristics, and biomolecular makeup. Exosomes, a subtype of EVs, are released by the fusion of multivesicular bodies (MVB) with the plasma membrane of the cell. Several methods have been described in literature to isolate exosomes from biofluids including blood, urine, milk, and cell culture media, among others. While differential ultracentrifugation (dUC) has been widely used to isolate exosomes, other techniques including ultrafiltration, precipitating agents such as poly-ethylene glycol (PEG), immunoaffinity capture, microfluidics, and size-exclusion chromatography (SEC) have emerged as credible alternatives with pros and cons associated with each. In this review, we provide a summary of commonly used exosomal isolation techniques with a focus on SEC as an ideal methodology. We evaluate the efficacy of SEC to isolate exosomes from an array of biological fluids, with a particular focus on its application to adipose tissue-derived exosomes. We argue that exosomes isolated via SEC are relatively pure and functional, and that this methodology is reproducible, scalable, inexpensive, and does not require specialized equipment or user expertise. However, it must be noted that while SEC is a good candidate method to isolate exosomes, direct comparative studies are required to support this conclusion.

scholarly journals Characterization of Extracellular Vesicles Labelled with a Lipophilic Dye Using Fluorescence Nanoparticle Tracking Analysis

Membranes ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 779
Author(s):  
Getnet Midekessa ◽  
Kasun Godakumara ◽  
Keerthie Dissanayake ◽  
Mohammad Mehedi Hasan ◽  
Qurat Ul Ain Reshi ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Fluorescent Dyes ◽  
Biological Fluid ◽  
Biological Fluids ◽  
Size Exclusion ◽  
Fluorescent Nanoparticles ◽  
Significant Shift ◽  
Reliable Technique ◽  
Exclusion Chromatography ◽  
Human Choriocarcinoma Cells

Research on extracellular vesicles (EVs) has intensified over the past decade, including fluorescent membrane labeling of EVs. An optimal fluorescent method requires the size of EVs to be preserved after labeling. Lipophilic fluorescent dyes, such as CellMask™ Green (CMG), have been widely used for this purpose. Here, we investigated conditions affecting the optimum CMG labeling of EVs derived from human choriocarcinoma cells (JAr) and different biological fluids using fluorescence NTA (fl-NTA). The effect of CMG labeling on the size, concentration and zeta potential (ZP) on JAr EVs purified with different methods were measured along with biological fluid-derived EVs. With the increase of CMG dye concentration, a significant decrease in the mean size of fluorescent nanoparticles (fl-NPs) was observed. The ZP of fl-NPs originating from JAr cells with the lowest and highest dye concentrations showed a significant shift towards more and less negative ZP values, respectively. Differences in the concentration of fl-NPs were observed for JAr EVs purified using size-exclusion chromatography (SEC) alone and SEC in combination with tangential flow filtration. The proportion of CMG labeling of NPs varied across different biological sources. CMG labeling may be a reliable technique for the detection of EVs using fl-NTA.

scholarly journals Micro size exclusion chromatography combined with a multiplex protein profiling method for extracellular vesicle protein detection from small sample volumes

2020 ◽  
Author(s):  
Li Sun ◽  
David Meckes
Keyword(s):  
Size Exclusion Chromatography ◽  
Biological Fluids ◽  
Protein Detection ◽  
Cancer Diagnostics ◽  
Small Sample ◽  
Surface Proteins ◽  
Protein Profiling ◽  
Size Exclusion ◽  
Exclusion Chromatography ◽  
Pcr Method

Abstract Extracellular vesicles (EVs) are small nanometer-sized membrane sacs secreted into biological fluids by all cells. EVs encapsulate proteins, RNAs and metabolites from its origin cell and play important roles in intercellular communication events. Over the past decade, EVs have become a new emerging source for cancer diagnostics. One of the challenges in the study of EVs and there utility as diagnostic biomarkers is the amount of EVs needed for traditional protein analysis methods. Here, we present a new immuno-PCR method that takes advantage of commercially available TotalSeq™ antibodies containing DNA conjugated oligos to identify immobilized protein analysts using real-time qPCR. Using this method, we demonstrate that multiple EV surface proteins can be profiled simultaneously with high sensitivity and specificity. This approach was also successfully applied to similar protocol using live cell and serum samples. We further described the development of a micro-size exclusion chromatography method, where we were able to detect EV surface proteins with as little as 10 μL of human serum when combined with immuno-PCR. Overall, these results show that the immuno-PCR method results in rapid detection of multiple EV markers from small sample volumes in a single tube.

scholarly journals A Review of Exosomal Isolation Methods: Is Size Exclusion Chromatography the Best Option?

2020 ◽  
Author(s):  
Karim Sidhom ◽  
Patience O. Obi ◽  
Ayesha Saleem
Keyword(s):  
Size Exclusion Chromatography ◽  
Culture Media ◽  
Biological Fluids ◽  
Vital Role ◽  
Size Exclusion ◽  
Isolation Techniques ◽  
Exclusion Chromatography ◽  
Specialized Equipment ◽  
Pros And Cons ◽  
Poly Ethylene

Extracellular vesicles (EVs) are membranous vesicles secreted by both prokaryotic and eukaryotic cells and play a vital role in intercellular communication. EVs are classified into several subtypes based on their origin, physical characteristics, and biomolecular makeup. Exosomes, a subtype of EVs, are released by the fusion of multivesicular bodies (MVB) with the plasma membrane of the cell. Several methods have been described in literature to isolate exosomes from biofluids including blood, urine, milk, and cell culture media among others. While differential ultracentrifugation (dUC), has been widely used to isolate exosomes, other techniques including ultrafiltration, precipitating agents such as poly-ethylene glycol (PEG), immunoaffinity capture, microfluidics and size exclusion chromatography (SEC) have emerged as credible alternatives with pros and cons associated with each. In this review, we provide a summary of commonly used exosomal isolation techniques with a focus on SEC as an ideal methodology. We evaluate the efficacy of SEC to isolate exosomes from an array of biological fluids, with a particular focus on its application to adipose tissue-derived exosomes. We argue that exosomes isolated via SEC are relatively pure and functional, and that this methodology is reproducible, scalable, inexpensive, and does not require specialized equipment or user expertise.

COMPARISONS OF NANOPARTICLE PROTEIN CORONA COMPLEXES ISOLATED WITH DIFFERENT METHODS

Nano LIFE ◽  
2013 ◽  
Vol 03 (04) ◽  
pp. 1343004 ◽  
Author(s):  
MARCO P. MONOPOLI ◽  
SHA WAN ◽  
FRANCESCA BALDELLI BOMBELLI ◽  
EUGENE MAHON ◽  
KENNETH A. DAWSON
Keyword(s):  
Size Exclusion Chromatography ◽  
Residence Time ◽  
Biological Fluids ◽  
Protein Composition ◽  
Protein Corona ◽  
Membrane Receptors ◽  
Size Exclusion ◽  
Dispersion Properties ◽  
Nanoparticle Surface ◽  
Exclusion Chromatography

Nanoparticles, after incubation in biological fluids, adsorb several kinds of biomolecules like lipids, sugars and mainly proteins with high affinities for the nanoparticle surface and with long residence time, forming the so-called hard corona. The biological machinery, such as cellular barriers and membrane receptors can directly engage with the protein corona while the pristine surface may remain inaccessible. Here we isolate nanoparticles associated with strongly bound biomolecules from the unbound and loosely bound ones, by different approaches: centrifugation, size exclusion chromatography and magnetic isolation. The different separation methodologies, despite requiring diverse time and operating mechanisms, gave nanoparticle-hard corona complexes which were found to be remarkably similar in both dispersion properties and protein composition thus proving to be equally valid.

Sign in / Sign up

Export Citation Format

Share Document