scholarly journals Isolation of Extracellular Vesicles from Biological Fluids via the Aggregation–Precipitation Approach for Downstream miRNAs Detection

Diagnostics ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 384
Author(s):  
Maria Y. Konoshenko ◽  
Evgeniy A. Lekchnov ◽  
Olga E. Bryzgunova ◽  
Elena Kiseleva ◽  
Inna A. Pyshnaya ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Biological Fluids ◽  
Low Cost ◽  
Precipitation Method ◽  
Medical Applications ◽  
Non Invasive ◽  
Clinical Laboratories ◽  
Healthy Donors ◽  
Invasive Diagnostics ◽  
Productive Method

Extracellular vesicles (EVs) have high potential as sources of biomarkers for non-invasive diagnostics. Thus, a simple and productive method of EV isolation is demanded for certain scientific and medical applications of EVs. Here we aim to develop a simple and effective method of EV isolation from different biofluids, suitable for both scientific, and clinical analyses of miRNAs transported by EVs. The proposed aggregation–precipitation method is based on the aggregation of EVs using dextran blue and the subsequent precipitation of EVs using 1.5% polyethylene glycol solutions. The developed method allows the effective isolation of EVs from plasma and urine. As shown using TEM, dynamic light scattering, and miRNA analyses, this method is not inferior to ultracentrifugation-based EV isolation in terms of its efficacy, lack of inhibitors for polymerase reactions and applicable for both healthy donors and cancer patients. This method is fast, simple, does not need complicated equipment, can be adapted for different biofluids, and has a low cost. The aggregation–precipitation method of EV isolation accessible and suitable for both research and clinical laboratories. This method has the potential to increase the diagnostic and prognostic utilization of EVs and miRNA-based diagnostics of urogenital pathologies.

scholarly journals Immunomagnetic sequential ultrafiltration (iSUF) platform for enrichment and purification of extracellular vesicles from biofluids

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Jingjing Zhang ◽  
Luong T. H. Nguyen ◽  
Richard Hickey ◽  
Nicole Walters ◽  
Xinyu Wang ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Culture Media ◽  
Metastatic Breast ◽  
Clinical Samples ◽  
Clinical Use ◽  
Immunomagnetic Beads ◽  
Liquid Biopsies ◽  
Cell Culture Media ◽  
Non Invasive ◽  
Healthy Donors

AbstractExtracellular vesicles (EVs) derived from tumor cells have the potential to provide a much-needed source of non-invasive molecular biomarkers for liquid biopsies. However, current methods for EV isolation have limited specificity towards tumor-derived EVs that limit their clinical use. Here, we present an approach called immunomagnetic sequential ultrafiltration (iSUF) that consists of sequential stages of purification and enrichment of EVs in approximately 2 h. In iSUF, EVs present in different volumes of biofluids (0.5–100 mL) can be significantly enriched (up to 1000 times), with up to 99% removal of contaminating proteins (e.g., albumin). The EV recovery rate by iSUF for cell culture media (CCM), serum, and urine corresponded to 98.0% ± 3.6%, 96.0% ± 2.0% and 94.0% ± 1.9%, respectively (p > 0.05). The final step of iSUF enables the separation of tumor-specific EVs by incorporating immunomagnetic beads to target EV subpopulations. Serum from a cohort of clinical samples from metastatic breast cancer (BC) patients and healthy donors were processed by the iSUF platform and the isolated EVs from patients showed significantly higher expression levels of BC biomarkers (i.e., HER2, CD24, and miR21).

scholarly journals Extracellular Vesicles-Based Biomarkers Represent a Promising Liquid Biopsy in Endometrial Cancer

Cancers ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 2000 ◽  
Author(s):  
Carolina Herrero ◽  
Alexandre de la Fuente ◽  
Carlos Casas-Arozamena ◽  
Victor Sebastian ◽  
Martin Prieto ◽  
...  
Keyword(s):  
Endometrial Cancer ◽  
Extracellular Vesicles ◽  
Biological Fluids ◽  
Low Cost ◽  
New Technology ◽  
Response To Therapy ◽  
Clinical Implementation ◽  
Clinical Value ◽  
Specific Content ◽  
Technical Performance

Tumor-derived extracellular vesicles (EVs) are secreted in large amounts into biological fluids of cancer patients. The analysis of EVs cargoes has been associated with patient´s outcome and response to therapy. However, current technologies for EVs isolation are tedious and low cost-efficient for routine clinical implementation. To explore the clinical value of circulating EVs analysis we attempted a proof-of-concept in endometrial cancer (EC) with ExoGAG, an easy to use and highly efficient new technology to enrich EVs. Technical performance was first evaluated using EVs secreted by Hec1A cells. Then, the clinical value of this strategy was questioned by analyzing the levels of two well-known tissue biomarkers in EC, L1 cell adhesion molecule (L1CAM) and Annexin A2 (ANXA2), in EVs purified from plasma in a cohort of 41 EC patients and 20 healthy controls. The results demonstrated the specific content of ANXA2 in the purified EVs fraction, with an accurate sensitivity and specificity for EC diagnosis. Importantly, high ANXA2 levels in circulating EVs were associated with high risk of recurrence and non-endometrioid histology suggesting a potential value as a prognostic biomarker in EC. These results also confirmed ExoGAG technology as a robust technique for the clinical implementation of circulating EVs analyses.

scholarly journals Calcium pyrophosphate powder derived from avian eggshell waste

Cerâmica ◽  
2016 ◽  
Vol 62 (363) ◽  
pp. 278-280 ◽  
Author(s):  
T. H. A. Corrêa ◽  
J. N. F. Holanda
Keyword(s):  
Electron Microscopy ◽  
Low Cost ◽  
Average Crystallite Size ◽  
Calcium Pyrophosphate ◽  
Precipitation Method ◽  
Medical Applications ◽  
X Ray Diffraction ◽  
X Ray ◽  
Eggshell Waste ◽  
Scanning Electron

Abstract Calcium pyrophosphate (CPP) was prepared by a simple precipitation method using avian eggshell waste as a low-cost alternative calcium precursor source. The synthesized CPP powder was characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM/EDS), thermogravimetric analysis (TGA), and Fourier transform infrared spectroscopy (FTIR). The results indicate that pure β-CPP nanocrystallites (Ca/P = 1.067) were successfully synthesized from avian eggshell waste. The correlation among XRD, SEM/EDS, TGA, and FTIR data is well established. The β-CPP particle exhibited spherical morphology with average crystallite size of 62.3 nm, and can be an important bioceramic for medical applications.

scholarly journals Immunomagnetic Sequential Ultrafiltration (iSUF) Platform for Enrichment and Purification of Extracellular Vesicles from Biofluids

2020 ◽  
Author(s):  
Jingjing Zhang ◽  
Luong TH Nguyen ◽  
Richard Hickey ◽  
Nicole Walters ◽  
Xinyu Wang ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Culture Media ◽  
Metastatic Breast ◽  
Clinical Samples ◽  
Immunomagnetic Beads ◽  
Liquid Biopsies ◽  
Cell Culture Media ◽  
Non Invasive ◽  
Healthy Donors ◽  
Small Cohort

AbstractExtracellular vesicles (EVs) derived from tumor cells have the potential to provide a much-needed source of non-invasive molecular biomarkers for liquid biopsies. However, current methods for EV isolation have limited specificity towards tumor-derived EVs that limit their clinical use. Here, we present an approach called immunomagnetic sequential ultrafiltration (iSUF) that consists of sequential stages of purification and enrichment of EVs (nonspecifically and specifically) in approximately 2 h. In iSUF, EVs present in different volumes of biofluids (0.5 mL to 100 mL) can be significantly enriched (up to 1000 times), with up to 99 % removal of contaminating proteins (e.g., albumin). The EV recovery rate by iSUF for cell culture media (CCM), serum, and urine corresponded to 98.0% ± 3.6%, 96.0% ± 2.0% and 94.0% ± 1.9%, respectively (p > 0.05). The final step of iSUF enables the separation of tumor-specific EVs by incorporating immunomagnetic beads specific to a target subpopulation of EVs. Serum from a small cohort of clinical samples from metastatic breast cancer (BC) patients and healthy donors were processed by the iSUF platform and the isolated EVs from patients showed significantly higher expression levels of BC biomarkers (i.e., HER2, CD24, and miR21).

scholarly journals Tomography for electrical impedance

2020 ◽  
Vol 16 (1) ◽  
Author(s):  
Juan David Muñoz Sánchez ◽  
Víctor Hugo Mosquera Leyton
Keyword(s):  
Electrical Impedance Tomography ◽  
Electrical Impedance ◽  
Wrist Joint ◽  
Low Cost ◽  
Good Alternative ◽  
Medical Applications ◽  
Joint Diseases ◽  
Impedance Tomography ◽  
Non Invasive ◽  
Joint Injuries

Introduction:This review article shows the state of the art of different techniques for monitoring joint injuries. This work is the product of the research project "Viability of electrical impedance tomography for the monitoring of joint injuries", which took place at the University of Cauca during the period 2018-2019. Aim:Identify non-invasive bio-image methods that are used in the evaluation of joint diseases. Methodology: Selection and review of papers related to the evaluation of joint injuries using non-invasive bio-image technologies using systematic mapping.  Results: Magnetic resonance and computed tomography systems make up the non-invasive methods of greater reliability and application in the evaluation of joint injuries. Similarly, some studies show good results from other methods such as systems based on bio-impedance when monitoring the deterioration of joint cartilage. However, electrical impedance tomography (EIT) devices have not yet been widely studied in the joint injuries evaluation. Conclusion: Electronic prototypes of low-cost electrical impedance tomography have been developed that have allowed for the detection and recognition of gestures made by hand from the analysis of the distribution of conductivity in the wrist joint, which allows us to infer that EIT could be a good alternative for the monitoring of joint injuries. Originality: The literature does not show studies focused on the development and implementation of EIT systems in medical applications related to joint injuries. Limitations: This review paper only mentions those studies that describe the non-invasive bio-image methods used to evaluate joint diseases, including the medical applications of EIT systems.

Separation of Extracellular Vesicles by DNA-Directed Immunocapturing Followed by Enzymatic Release

2020 ◽  
Author(s):  
Dario Brambilla ◽  
Laura Sola ◽  
Elisa Chiodi ◽  
Natasa Zarovni ◽  
Diogo Fortunato ◽  
...  
Keyword(s):  
Cell Culture ◽  
Extracellular Vesicles ◽  
Culture Media ◽  
Biological Fluids ◽  
Imaging Techniques ◽  
Cell Communication ◽  
Disease Diagnosis ◽  
Cell Culture Supernatant ◽  
Transmission Electron ◽  
Downstream Analysis

Extracellular vesicles (EVs) have attracted great interest among researchers due to their role in cell-cell communication, disease diagnosis, and drug delivery. In spite of their potential in the medical field, there is no consensus on the best method for separating microvesicles from cell culture supernatant and complex biological fluids. Obtaining a good recovery yield and preserving physical characteristics is critical for the diagnostic and therapeutic use of EVs. The separation is made complex by the fact that blood and cell culture media, contain a large number of nanoparticles in the same size range. Methods that exploit immunoaffinity capture provide high purity samples and overcome the issues of currently used separation methods. However, the release of captured nanovesicles requires harsh conditions that hinder their use in certain types of downstream analysis. Herein, a novel capture and release approach for small extracellular vesicles (sEVs), based on DNAdirected immobilization of antiCD63 antibody is presented. The flexible DNAlinker increases the capture efficiency and allows releasing of EVs by exploiting the endonucleasic activity of DNAse I. This separation protocol works under mild conditions, enabling the release of intact vesicles that can be successfully analyzed by imaging techniques. In this article sEVs recovered from plasma were characterized by established techniques for EVs analysis including nanoparticle tracking and transmission electron microscopy.<br>

Microextraction and Chromatographic Analysis of Budesonide Epimers in Exhaled Breath Condensate

2020 ◽  
Vol 16 (8) ◽  
pp. 1032-1040
Author(s):  
Laleh Samini ◽  
Maryam Khoubnasabjafari ◽  
Mohamad M. Alimorad ◽  
Vahid Jouyban-Gharamaleki ◽  
Hak-Kim Chan ◽  
...  
Keyword(s):  
Biological Fluids ◽  
Exhaled Breath Condensate ◽  
Low Cost ◽  
Exhaled Breath ◽  
Chromatography Method ◽  
Extraction Solvent ◽  
Drug Concentrations ◽  
Dispersive Liquid Liquid Microextraction ◽  
Breath Condensate

Background: Analysis of drug concentrations in biological fluids is required in clinical sciences for various purposes. Among other biological samples, exhaled breath condensate (EBC) is a potential sample for follow up of drug concentrations. Methods: A dispersive liquid-liquid microextraction (DLLME) procedure followed by a validated liquid chromatography method was employed for the determination of budesonide (BDS) in EBC samples collected using a homemade setup. EBC is a non-invasive biological sample with possible applications for monitoring drug concentrations. The proposed analytical method is validated according to the FDA guidelines using EBC-spiked samples. Its applicability is tested on EBC samples collected from healthy volunteers receiving a single puff of BDS. Results: The best DLLME conditions involved the use of methanol (1 mL) as a disperser solvent, chloroform (200 μL) as an extraction solvent, and centrifugation rate of 3500 rpm for 5 minutes. The method was validated over a concentration range of 21-210 μg·L-1 in EBC. Inter- and intra-day precisions were less than 10% where the acceptable levels are less than 20%. The validated method was successfully applied for the determination of BDS in EBC samples. Conclusion: The findings of this study indicate that the developed method can be used for the extraction and quantification of BDS in EBC samples using a low cost method.

scholarly journals Non-invasive diagnostics in fossils - Magnetic Resonance Imaging of pathological belemnites

2005 ◽  
Vol 2 (2) ◽  
pp. 133-140 ◽  
Author(s):  
D. Mietchen ◽  
H. Keupp ◽  
B. Manz ◽  
F. Volke
Keyword(s):  
Magnetic Resonance Imaging ◽  
Magnetic Resonance ◽  
Soft Tissue ◽  
Clinical Diagnostics ◽  
Resonance Imaging ◽  
Non Invasive ◽  
Physiological Processes ◽  
Magnetic Resonance Imaging Mri ◽  
Invasive Diagnostics

Abstract. For more than a decade, Magnetic Resonance Imaging (MRI) has been routinely employed in clinical diagnostics because it allows non-invasive studies of anatomical structures and physiological processes in vivo and to differentiate between healthy and pathological states, particularly of soft tissue. Here, we demonstrate that MRI can likewise be applied to fossilized biological samples and help in elucidating paleopathological and paleoecological questions: Five anomalous guards of Jurassic and Cretaceous belemnites are presented along with putative paleopathological diagnoses directly derived from 3D MR images with microscopic resolution. Syn vivo deformities of both the mineralized internal rostrum and the surrounding former soft tissue can be traced back in part to traumatic events of predator-prey-interactions, and partly to parasitism. Besides, evidence is presented that the frequently observed anomalous apical collar might be indicative of an inflammatory disease. These findings highlight the potential of Magnetic Resonance techniques for further paleontological applications.

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