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  1. Article ; Online: Multi-marker profiling of extracellular vesicles using streaming current and sequential electrostatic labeling.

    Sahu, Siddharth S / Gevari, Moein T / Nagy, Ábel / Gestin, Maxime / Hååg, Petra / Lewensohn, Rolf / Viktorsson, Kristina / Karlström, Amelie E / Dev, Apurba

    Biosensors & bioelectronics

    2023  Volume 227, Page(s) 115142

    Abstract: High heterogeneity in the membrane protein expression of small extracellular vesicles (sEVs) means that bulk methods relying on antibody-based capture for expression analysis have a drawback that each type of antibody may capture a different sub- ... ...

    Abstract High heterogeneity in the membrane protein expression of small extracellular vesicles (sEVs) means that bulk methods relying on antibody-based capture for expression analysis have a drawback that each type of antibody may capture a different sub-population. An improved approach is to capture a representative sEV population, without any bias, and then perform a multiplexed protein expression analysis on this population. However, such a possibility has been largely limited to fluorescence-based methods. Here, we present a novel electrostatic labelling strategy and a microchip-based all-electric method for membrane protein analysis of sEVs. The method allows us to profile multiple surface proteins on the captured sEVs using alternating charge labels. It also permits the comparison of expression levels in different sEV-subtypes. The proof of concept was tested by capturing sEVs both non-specifically (unbiased) as well as via anti-CD9 capture probes (biased), and then profiling the expression levels of various surface proteins using the charge labelled antibodies. The method is the first of its kind, demonstrating an all-electrical and microchip based method that allows for unbiased analysis of sEV membrane protein expression, comparison of expression levels in different sEV subsets, and fractional estimation of different sEV sub-populations. These results were also validated in parallel using a single-sEV fluorescence technique.
    MeSH term(s) Static Electricity ; Biosensing Techniques ; Extracellular Vesicles ; Electricity ; Antibodies ; Membrane Proteins
    Chemical Substances Antibodies ; Membrane Proteins
    Language English
    Publishing date 2023-02-09
    Publishing country England
    Document type Journal Article
    ZDB-ID 1011023-9
    ISSN 1873-4235 ; 0956-5663
    ISSN (online) 1873-4235
    ISSN 0956-5663
    DOI 10.1016/j.bios.2023.115142
    Database MEDical Literature Analysis and Retrieval System OnLINE

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    In stock of ZB MED Cologne/Königswinter

    Zs.A 2275: Show issues Location:
    Je nach Verfügbarkeit (siehe Angabe bei Bestand)
    bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular
    Jg. 1995 - 2021: Lesesall (2.OG)
    ab Jg. 2022: Lesesaal (EG)

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  2. Article ; Online: Multi-marker profiling of extracellular vesicles using streaming current and sequential electrostatic labeling

    Sahu, Siddharth S. / Gevari, Moein T. / Nagy, Ábel / Gestin, Maxime / Haag, Petra / Lewensohn, Rolf / Viktorsson, Kristina / Karlström, Amelie E. / Dev, Apurba

    Biosensors and Bioelectronics. 2023 Feb. 09, p.115142-

    2023  , Page(s) 115142–

    Abstract: High heterogeneity in the membrane protein expression of small extracellular vesicles (sEVs) means that bulk methods relying on antibody-based capture for expression analysis have a drawback that each type of antibody may capture a different sub- ... ...

    Abstract High heterogeneity in the membrane protein expression of small extracellular vesicles (sEVs) means that bulk methods relying on antibody-based capture for expression analysis have a drawback that each type of antibody may capture a different sub-population. An improved approach is to capture a representative sEV population, without any bias, and then perform a multiplexed protein expression analysis on this population. However, such a possibility has been largely limited to fluorescence-based methods. Here, we present a novel electrostatic labelling strategy and a microchip-based all-electric method for membrane protein analysis of sEVs. The method allows us to profile multiple surface proteins on the captured sEVs using alternating charge labels. It also permits the comparison of expression levels in different sEV-subtypes. The proof of concept was tested by capturing sEVs both non-specifically (unbiased) as well as via anti-CD9 capture probes (biased), and then profiling the expression levels of various surface proteins using the charge labelled antibodies. The method is the first of its kind, demonstrating an all-electrical and microchip based method that allows for unbiased analysis of sEV membrane protein expression, comparison of expression levels in different sEV subsets, and fractional estimation of different sEV sub-populations. These results were also validated in parallel using a single-sEV fluorescence technique.
    Keywords antibodies ; biosensors ; fluorescence ; membrane proteins ; microchip technology ; protein synthesis ; Extracellular vesicles ; Streaming current ; Electrostatic labels
    Language English
    Dates of publication 2023-0209
    Publishing place Elsevier B.V.
    Document type Article ; Online
    Note Pre-press version
    ZDB-ID 1011023-9
    ISSN 1873-4235 ; 0956-5663
    ISSN (online) 1873-4235
    ISSN 0956-5663
    DOI 10.1016/j.bios.2023.115142
    Database NAL-Catalogue (AGRICOLA)

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    In stock of ZB MED Cologne/Königswinter

    Zs.A 2275: Show issues Location:
    Je nach Verfügbarkeit (siehe Angabe bei Bestand)
    bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular
    Jg. 1995 - 2021: Lesesall (2.OG)
    ab Jg. 2022: Lesesaal (EG)

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  3. Article: Multiplexed electrokinetic sensor for detection and therapy monitoring of extracellular vesicles from liquid biopsies of non-small-cell lung cancer patients

    Cavallaro, Sara / Hååg, Petra / Sahu, Siddharth S. / Berisha, Lorenca / Kaminskyy, Vitaliy O. / Ekman, Simon / Lewensohn, Rolf / Linnros, Jan / Viktorsson, Kristina / Dev, Apurba

    Biosensors & bioelectronics. 2021 Dec. 01, v. 193

    2021  

    Abstract: Liquid biopsies based on extracellular vesicles (EVs) represent a promising tool for treatment monitoring of tumors, including non-small-cell lung cancers (NSCLC). In this study, we report on a multiplexed electrokinetic sensor for surface protein ... ...

    Abstract Liquid biopsies based on extracellular vesicles (EVs) represent a promising tool for treatment monitoring of tumors, including non-small-cell lung cancers (NSCLC). In this study, we report on a multiplexed electrokinetic sensor for surface protein profiling of EVs from clinical samples. The method detects the difference in the streaming current generated by EV binding to the surface of a functionalized microcapillary, thereby estimating the expression level of a marker. Using multiple microchannels functionalized with different antibodies in a parallel fluidic connection, we first demonstrate the capacity for simultaneous detection of multiple surface markers in small EVs (sEVs) from NSCLC cells. To investigate the prospects of liquid biopsies based on EVs, we then apply the method to profile sEVs isolated from the pleural effusion (PE) fluids of five NSCLC patients with different genomic alterations (ALK, KRAS or EGFR) and applied treatments (chemotherapy, EGFR- or ALK-tyrosine kinase inhibitors). The vesicles were targeted against CD9, as well as EGFR and PD-L1, two treatment targets in NSCLC. The electrokinetic signals show detection of these markers on sEVs, highlighting distinct interpatient differences, e.g., increased EGFR levels in sEVs from a patient with EGFR mutation as compared to an ALK-fusion one. The sensors also detect differences in PD-L1 expressions. The analysis of sEVs from a patient prior and post ALK-TKI crizotinib treatment reveals significant increases in the expressions of some markers (EGFR and PD-L1). These results hold promise for the application of the method for tumor treatment monitoring based on sEVs from patient liquid biopsies.
    Keywords biosensors ; drug therapy ; genomics ; liquids ; lung neoplasms ; lungs ; mutation ; patients ; surface proteins
    Language English
    Dates of publication 2021-1201
    Publishing place Elsevier B.V.
    Document type Article
    ZDB-ID 1011023-9
    ISSN 1873-4235 ; 0956-5663
    ISSN (online) 1873-4235
    ISSN 0956-5663
    DOI 10.1016/j.bios.2021.113568
    Database NAL-Catalogue (AGRICOLA)

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    In stock of ZB MED Cologne/Königswinter

    Zs.A 2275: Show issues Location:
    Je nach Verfügbarkeit (siehe Angabe bei Bestand)
    bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular
    Jg. 1995 - 2021: Lesesall (2.OG)
    ab Jg. 2022: Lesesaal (EG)

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  4. Article ; Online: Multiplexed electrokinetic sensor for detection and therapy monitoring of extracellular vesicles from liquid biopsies of non-small-cell lung cancer patients.

    Cavallaro, Sara / Hååg, Petra / Sahu, Siddharth S / Berisha, Lorenca / Kaminskyy, Vitaliy O / Ekman, Simon / Lewensohn, Rolf / Linnros, Jan / Viktorsson, Kristina / Dev, Apurba

    Biosensors & bioelectronics

    2021  Volume 193, Page(s) 113568

    Abstract: Liquid biopsies based on extracellular vesicles (EVs) represent a promising tool for treatment monitoring of tumors, including non-small-cell lung cancers (NSCLC). In this study, we report on a multiplexed electrokinetic sensor for surface protein ... ...

    Abstract Liquid biopsies based on extracellular vesicles (EVs) represent a promising tool for treatment monitoring of tumors, including non-small-cell lung cancers (NSCLC). In this study, we report on a multiplexed electrokinetic sensor for surface protein profiling of EVs from clinical samples. The method detects the difference in the streaming current generated by EV binding to the surface of a functionalized microcapillary, thereby estimating the expression level of a marker. Using multiple microchannels functionalized with different antibodies in a parallel fluidic connection, we first demonstrate the capacity for simultaneous detection of multiple surface markers in small EVs (sEVs) from NSCLC cells. To investigate the prospects of liquid biopsies based on EVs, we then apply the method to profile sEVs isolated from the pleural effusion (PE) fluids of five NSCLC patients with different genomic alterations (ALK, KRAS or EGFR) and applied treatments (chemotherapy, EGFR- or ALK-tyrosine kinase inhibitors). The vesicles were targeted against CD9, as well as EGFR and PD-L1, two treatment targets in NSCLC. The electrokinetic signals show detection of these markers on sEVs, highlighting distinct interpatient differences, e.g., increased EGFR levels in sEVs from a patient with EGFR mutation as compared to an ALK-fusion one. The sensors also detect differences in PD-L1 expressions. The analysis of sEVs from a patient prior and post ALK-TKI crizotinib treatment reveals significant increases in the expressions of some markers (EGFR and PD-L1). These results hold promise for the application of the method for tumor treatment monitoring based on sEVs from patient liquid biopsies.
    MeSH term(s) Anaplastic Lymphoma Kinase/genetics ; Biosensing Techniques ; Carcinoma, Non-Small-Cell Lung/drug therapy ; ErbB Receptors/genetics ; Extracellular Vesicles ; Humans ; Liquid Biopsy ; Lung Neoplasms/diagnosis ; Lung Neoplasms/drug therapy ; Mutation ; Protein Kinase Inhibitors/therapeutic use
    Chemical Substances Protein Kinase Inhibitors ; Anaplastic Lymphoma Kinase (EC 2.7.10.1) ; ErbB Receptors (EC 2.7.10.1)
    Language English
    Publishing date 2021-08-17
    Publishing country England
    Document type Journal Article
    ZDB-ID 1011023-9
    ISSN 1873-4235 ; 0956-5663
    ISSN (online) 1873-4235
    ISSN 0956-5663
    DOI 10.1016/j.bios.2021.113568
    Database MEDical Literature Analysis and Retrieval System OnLINE

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    Kategorien

    In stock of ZB MED Cologne/Königswinter

    Zs.A 2275: Show issues Location:
    Je nach Verfügbarkeit (siehe Angabe bei Bestand)
    bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular
    Jg. 1995 - 2021: Lesesall (2.OG)
    ab Jg. 2022: Lesesaal (EG)

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  5. Article ; Online: Multiparametric Profiling of Single Nanoscale Extracellular Vesicles by Combined Atomic Force and Fluorescence Microscopy: Correlation and Heterogeneity in Their Molecular and Biophysical Features.

    Cavallaro, Sara / Pevere, Federico / Stridfeldt, Fredrik / Görgens, André / Paba, Carolina / Sahu, Siddharth S / Mamand, Doste R / Gupta, Dhanu / El Andaloussi, Samir / Linnros, Jan / Dev, Apurba

    Small (Weinheim an der Bergstrasse, Germany)

    2021  Volume 17, Issue 14, Page(s) e2008155

    Abstract: Being a key player in intercellular communications, nanoscale extracellular vesicles (EVs) offer unique opportunities for both diagnostics and therapeutics. However, their cellular origin and functional identity remain elusive due to the high ... ...

    Abstract Being a key player in intercellular communications, nanoscale extracellular vesicles (EVs) offer unique opportunities for both diagnostics and therapeutics. However, their cellular origin and functional identity remain elusive due to the high heterogeneity in their molecular and physical features. Here, for the first time, multiple EV parameters involving membrane protein composition, size and mechanical properties on single small EVs (sEVs) are simultaneously studied by combined fluorescence and atomic force microscopy. Furthermore, their correlation and heterogeneity in different cellular sources are investigated. The study, performed on sEVs derived from human embryonic kidney 293, cord blood mesenchymal stromal and human acute monocytic leukemia cell lines, identifies both common and cell line-specific sEV subpopulations bearing distinct distributions of the common tetraspanins (CD9, CD63, and CD81) and biophysical properties. Although the tetraspanin abundances of individual sEVs are independent of their sizes, the expression levels of CD9 and CD63 are strongly correlated. A sEV population co-expressing all the three tetraspanins in relatively high abundance, however, having average diameters of <100 nm and relatively low Young moduli, is also found in all cell lines. Such a multiparametric approach is expected to provide new insights regarding EV biology and functions, potentially deciphering unsolved questions in this field.
    MeSH term(s) Biophysics ; Cell Communication ; Child ; Extracellular Vesicles ; Humans ; Microscopy, Fluorescence ; Tetraspanins
    Chemical Substances Tetraspanins
    Language English
    Publishing date 2021-03-07
    Publishing country Germany
    Document type Journal Article ; Research Support, Non-U.S. Gov't ; Review
    ZDB-ID 2168935-0
    ISSN 1613-6829 ; 1613-6810
    ISSN (online) 1613-6829
    ISSN 1613-6810
    DOI 10.1002/smll.202008155
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  6. Article ; Online: Label-Free Surface Protein Profiling of Extracellular Vesicles by an Electrokinetic Sensor.

    Cavallaro, Sara / Horak, Josef / Hååg, Petra / Gupta, Dhanu / Stiller, Christiane / Sahu, Siddharth S / Görgens, André / Gatty, Hithesh K / Viktorsson, Kristina / El Andaloussi, Samir / Lewensohn, Rolf / Karlström, Amelie E / Linnros, Jan / Dev, Apurba

    ACS sensors

    2019  Volume 4, Issue 5, Page(s) 1399–1408

    Abstract: Small extracellular vesicles (sEVs) generated from the endolysosomal system, often referred to as exosomes, have attracted interest as a suitable biomarker for cancer diagnostics, as they carry valuable biological information and reflect their cells of ... ...

    Abstract Small extracellular vesicles (sEVs) generated from the endolysosomal system, often referred to as exosomes, have attracted interest as a suitable biomarker for cancer diagnostics, as they carry valuable biological information and reflect their cells of origin. Herein, we propose a simple and inexpensive electrical method for label-free detection and profiling of sEVs in the size range of exosomes. The detection method is based on the electrokinetic principle, where the change in the streaming current is monitored as the surface markers of the sEVs interact with the affinity reagents immobilized on the inner surface of a silica microcapillary. As a proof-of-concept, we detected sEVs derived from the non-small-cell lung cancer (NSCLC) cell line H1975 for a set of representative surface markers, such as epidermal growth factor receptor (EGFR), CD9, and CD63. The detection sensitivity was estimated to be ∼175000 sEVs, which represents a sensor surface coverage of only 0.04%. We further validated the ability of the sensor to measure the expression level of a membrane protein by using sEVs displaying artificially altered expressions of EGFR and CD63, which were derived from NSCLC and human embryonic kidney (HEK) 293T cells, respectively. The analysis revealed that the changes in EGFR and CD63 expressions in sEVs can be detected with a sensitivity in the order of 10% and 3%, respectively, of their parental cell expressions. The method can be easily parallelized and combined with existing microfluidic-based EV isolation technologies, allowing for rapid detection and monitoring of sEVs for cancer diagnosis.
    MeSH term(s) Biomarkers/metabolism ; Cell Line, Tumor ; Electric Conductivity ; ErbB Receptors/metabolism ; Extracellular Vesicles/metabolism ; HEK293 Cells ; Humans ; Tetraspanin 30/metabolism
    Chemical Substances Biomarkers ; Tetraspanin 30 ; EGFR protein, human (EC 2.7.10.1) ; ErbB Receptors (EC 2.7.10.1)
    Language English
    Publishing date 2019-05-07
    Publishing country United States
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ISSN 2379-3694
    ISSN (online) 2379-3694
    DOI 10.1021/acssensors.9b00418
    Database MEDical Literature Analysis and Retrieval System OnLINE

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