University of Hertfordshire

By the same authors

Real time analysis of microvesiculation using a Quartz Crystal Microbalance

Research output: Contribution to conferenceAbstractpeer-review

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Real time analysis of microvesiculation using a Quartz Crystal Microbalance. / Inal, Jameel; Stratton, Dan.

2014. 37-37 Abstract from Third International Meeting of ISEV 2014: Rotterdam, The Netherlands, April 30th–May 3rd, 2014, Rotterdam, Netherlands.

Research output: Contribution to conferenceAbstractpeer-review

Harvard

Inal, J & Stratton, D 2014, 'Real time analysis of microvesiculation using a Quartz Crystal Microbalance', Third International Meeting of ISEV 2014: Rotterdam, The Netherlands, April 30th–May 3rd, 2014, Rotterdam, Netherlands, 30/04/14 - 3/05/14 pp. 37-37. https://doi.org/10.3402/jev.v3.24214

APA

Inal, J., & Stratton, D. (2014). Real time analysis of microvesiculation using a Quartz Crystal Microbalance. 37-37. Abstract from Third International Meeting of ISEV 2014: Rotterdam, The Netherlands, April 30th–May 3rd, 2014, Rotterdam, Netherlands. https://doi.org/10.3402/jev.v3.24214

Vancouver

Inal J, Stratton D. Real time analysis of microvesiculation using a Quartz Crystal Microbalance. 2014. Abstract from Third International Meeting of ISEV 2014: Rotterdam, The Netherlands, April 30th–May 3rd, 2014, Rotterdam, Netherlands. https://doi.org/10.3402/jev.v3.24214

Author

Inal, Jameel ; Stratton, Dan. / Real time analysis of microvesiculation using a Quartz Crystal Microbalance. Abstract from Third International Meeting of ISEV 2014: Rotterdam, The Netherlands, April 30th–May 3rd, 2014, Rotterdam, Netherlands.1 p.

Bibtex

@conference{3a0594bf64cb4f1d8a7b3d8ae12198ef,
title = "Real time analysis of microvesiculation using a Quartz Crystal Microbalance",
abstract = "Introduction: Characterization of microvesicles (MVs) is essential forunderstanding their mechanisms of action and biological importance.Stimulated MVs (sMVs) are released through the activation of cells bya multitude of factors. We aimed to use a quartz crystal microbalance(QCM) or piezoelectric quartz resonator able to determine small masschanges, to monitor MV release and to determine MV mass. Methods:A QCM (Q-Sence E1) was used to analyse MV release from THP-1leukaemic promonocytes. The cells in RPMI and 2 mM Ca2_ wereapplied to the QCM to establish a steady baseline. The sample on thesensor was stimulated to microvesiculate with 10% exosome- andMV-free normal human serum. The QCM was then able to monitorsample density and fluid rigidity. Over the same time frame, the levelof apoptosis of cells releasing MVs was assessed by staining withannexin V and 7-aminoactinomycin D (Guava Nexin Reagent). Usingthe QCM we were also able to measure MV mass directly bymeasuring their ability to quench the oscillating momentum of theQCM. Results: Using the QCM, we were able to monitor deposition ofcells on the crystal and then sMV release from cells, in the absence ofany labelling or fluorescent probe, by measuring cell mass change.Cells (105) were deposited onto the QCM electrodes, and thefrequency decreases over the first 1000s indicating attachment. Thecells were then stimulated with 10% EV-free NHS in RPMI and Ca2_(2 mM) or, as a control, with heat inactivated NHS. During the ensuing6.5 min, the resonate frequency remained stable. Then, over thefollowing 10 min there was a 30 Hz increase indicating a loss in mass,consistent with the high rate of sMV observed. Given the crystalconstant, C as 17.7, ^f as 19 Hz and v (the third overtone) as 3, andwith the crystal area at 0.2 cm2, using the Sauerbrey equation wecalculated the mass loss to be 23 ng which corresponded to 0.25 pgper MV given that 0.92 _ 105 MVs were released. The 16 min periodover which MVs continue to be released as determined on the QCMcoincides with the MV increase measured by FACS and with anincrease in early apoptosis from 4% plateauing at 10%, levels oflate apoptosis remaining at 1_3%. We also looked at deposition ofsMV on the sensor. Given a Df of 27197 Hz for the depositionof 1.3_106 sMVs, we estimate the mass of an sMV by this approachas 0.24190.006 pg. Summary/conclusion: Using the QCM we wereable to measure a significant change in cellular mass, beginning at 6.5min post-stimulus and peaking at 1000 s post-stimulus. The QCM alsodetected a decrease in media fluidity, attributed to the process ofmembrane blebbing on THP-1 and MV release. The QCM was ableto provide an accurate measurement of sMV mass (0.25 pg) bycalculating the loss in mass of the stimulated cells. By measuring thequenching of the oscillating momentum on the QCM as sMVs aredeposited on the sensor, we were also able to calculate the mass ofan sMV as 0.24 pg.",
author = "Jameel Inal and Dan Stratton",
year = "2014",
month = apr,
day = "25",
doi = "10.3402/jev.v3.24214",
language = "English",
pages = "37--37",
note = "Third International Meeting of ISEV 2014: Rotterdam, The Netherlands, April 30th–May 3rd, 2014 : International Society for Extracellular Vesicles annual conference, ISEV, 2014 ; Conference date: 30-04-2014 Through 03-05-2014",
url = "http://www.tandfonline.com/doi/abs/10.3402/jev.v3.24214",

}

RIS

TY - CONF

T1 - Real time analysis of microvesiculation using a Quartz Crystal Microbalance

AU - Inal, Jameel

AU - Stratton, Dan

PY - 2014/4/25

Y1 - 2014/4/25

N2 - Introduction: Characterization of microvesicles (MVs) is essential forunderstanding their mechanisms of action and biological importance.Stimulated MVs (sMVs) are released through the activation of cells bya multitude of factors. We aimed to use a quartz crystal microbalance(QCM) or piezoelectric quartz resonator able to determine small masschanges, to monitor MV release and to determine MV mass. Methods:A QCM (Q-Sence E1) was used to analyse MV release from THP-1leukaemic promonocytes. The cells in RPMI and 2 mM Ca2_ wereapplied to the QCM to establish a steady baseline. The sample on thesensor was stimulated to microvesiculate with 10% exosome- andMV-free normal human serum. The QCM was then able to monitorsample density and fluid rigidity. Over the same time frame, the levelof apoptosis of cells releasing MVs was assessed by staining withannexin V and 7-aminoactinomycin D (Guava Nexin Reagent). Usingthe QCM we were also able to measure MV mass directly bymeasuring their ability to quench the oscillating momentum of theQCM. Results: Using the QCM, we were able to monitor deposition ofcells on the crystal and then sMV release from cells, in the absence ofany labelling or fluorescent probe, by measuring cell mass change.Cells (105) were deposited onto the QCM electrodes, and thefrequency decreases over the first 1000s indicating attachment. Thecells were then stimulated with 10% EV-free NHS in RPMI and Ca2_(2 mM) or, as a control, with heat inactivated NHS. During the ensuing6.5 min, the resonate frequency remained stable. Then, over thefollowing 10 min there was a 30 Hz increase indicating a loss in mass,consistent with the high rate of sMV observed. Given the crystalconstant, C as 17.7, ^f as 19 Hz and v (the third overtone) as 3, andwith the crystal area at 0.2 cm2, using the Sauerbrey equation wecalculated the mass loss to be 23 ng which corresponded to 0.25 pgper MV given that 0.92 _ 105 MVs were released. The 16 min periodover which MVs continue to be released as determined on the QCMcoincides with the MV increase measured by FACS and with anincrease in early apoptosis from 4% plateauing at 10%, levels oflate apoptosis remaining at 1_3%. We also looked at deposition ofsMV on the sensor. Given a Df of 27197 Hz for the depositionof 1.3_106 sMVs, we estimate the mass of an sMV by this approachas 0.24190.006 pg. Summary/conclusion: Using the QCM we wereable to measure a significant change in cellular mass, beginning at 6.5min post-stimulus and peaking at 1000 s post-stimulus. The QCM alsodetected a decrease in media fluidity, attributed to the process ofmembrane blebbing on THP-1 and MV release. The QCM was ableto provide an accurate measurement of sMV mass (0.25 pg) bycalculating the loss in mass of the stimulated cells. By measuring thequenching of the oscillating momentum on the QCM as sMVs aredeposited on the sensor, we were also able to calculate the mass ofan sMV as 0.24 pg.

AB - Introduction: Characterization of microvesicles (MVs) is essential forunderstanding their mechanisms of action and biological importance.Stimulated MVs (sMVs) are released through the activation of cells bya multitude of factors. We aimed to use a quartz crystal microbalance(QCM) or piezoelectric quartz resonator able to determine small masschanges, to monitor MV release and to determine MV mass. Methods:A QCM (Q-Sence E1) was used to analyse MV release from THP-1leukaemic promonocytes. The cells in RPMI and 2 mM Ca2_ wereapplied to the QCM to establish a steady baseline. The sample on thesensor was stimulated to microvesiculate with 10% exosome- andMV-free normal human serum. The QCM was then able to monitorsample density and fluid rigidity. Over the same time frame, the levelof apoptosis of cells releasing MVs was assessed by staining withannexin V and 7-aminoactinomycin D (Guava Nexin Reagent). Usingthe QCM we were also able to measure MV mass directly bymeasuring their ability to quench the oscillating momentum of theQCM. Results: Using the QCM, we were able to monitor deposition ofcells on the crystal and then sMV release from cells, in the absence ofany labelling or fluorescent probe, by measuring cell mass change.Cells (105) were deposited onto the QCM electrodes, and thefrequency decreases over the first 1000s indicating attachment. Thecells were then stimulated with 10% EV-free NHS in RPMI and Ca2_(2 mM) or, as a control, with heat inactivated NHS. During the ensuing6.5 min, the resonate frequency remained stable. Then, over thefollowing 10 min there was a 30 Hz increase indicating a loss in mass,consistent with the high rate of sMV observed. Given the crystalconstant, C as 17.7, ^f as 19 Hz and v (the third overtone) as 3, andwith the crystal area at 0.2 cm2, using the Sauerbrey equation wecalculated the mass loss to be 23 ng which corresponded to 0.25 pgper MV given that 0.92 _ 105 MVs were released. The 16 min periodover which MVs continue to be released as determined on the QCMcoincides with the MV increase measured by FACS and with anincrease in early apoptosis from 4% plateauing at 10%, levels oflate apoptosis remaining at 1_3%. We also looked at deposition ofsMV on the sensor. Given a Df of 27197 Hz for the depositionof 1.3_106 sMVs, we estimate the mass of an sMV by this approachas 0.24190.006 pg. Summary/conclusion: Using the QCM we wereable to measure a significant change in cellular mass, beginning at 6.5min post-stimulus and peaking at 1000 s post-stimulus. The QCM alsodetected a decrease in media fluidity, attributed to the process ofmembrane blebbing on THP-1 and MV release. The QCM was ableto provide an accurate measurement of sMV mass (0.25 pg) bycalculating the loss in mass of the stimulated cells. By measuring thequenching of the oscillating momentum on the QCM as sMVs aredeposited on the sensor, we were also able to calculate the mass ofan sMV as 0.24 pg.

U2 - 10.3402/jev.v3.24214

DO - 10.3402/jev.v3.24214

M3 - Abstract

SP - 37

EP - 37

T2 - Third International Meeting of ISEV 2014: Rotterdam, The Netherlands, April 30th–May 3rd, 2014

Y2 - 30 April 2014 through 3 May 2014

ER -