Profiling the iron, copper and zinc content in primary neuron and astrocyte cultures by rapid online quantitative size exclusion chromatography-inductively coupled plasma-mass spectrometry

Hare, DJ; Grubman, A; Ryan, TM; Lothian, A; Liddell, JR; Grimm, R; Matsuda, T; Doble, PA; Cherny, RA; Bush, AI; White, AR; Masters, CL; Roberts, BR

Profiling the iron, copper and zinc content in primary neuron and astrocyte cultures by rapid online quantitative size exclusion chromatography-inductively coupled plasma-mass spectrometry

Hare, DJ

Grubman, A Ryan, TM Lothian, A Liddell, JR Grimm, R Matsuda, T Doble, PA

Cherny, RA Bush, AI White, AR Masters, CL Roberts, BR

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Publication Type:: Journal Article
Citation:: Metallomics, 2013, 5 (12), pp. 1656 - 1662
Issue Date:: 2013-12-01

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Field	Value	Language
dc.contributor.author	Hare, DJ https://orcid.org/0000-0002-5922-7643	en_US
dc.contributor.author	Grubman, A	en_US
dc.contributor.author	Ryan, TM	en_US
dc.contributor.author	Lothian, A	en_US
dc.contributor.author	Liddell, JR	en_US
dc.contributor.author	Grimm, R	en_US
dc.contributor.author	Matsuda, T	en_US
dc.contributor.author	Doble, PA https://orcid.org/0000-0002-8472-1301	en_US
dc.contributor.author	Cherny, RA	en_US
dc.contributor.author	Bush, AI	en_US
dc.contributor.author	White, AR	en_US
dc.contributor.author	Masters, CL	en_US
dc.contributor.author	Roberts, BR	en_US
dc.date.issued	2013-12-01	en_US
dc.identifier.citation	Metallomics, 2013, 5 (12), pp. 1656 - 1662	en_US
dc.identifier.issn	1756-5901	en_US
dc.identifier.uri	http://hdl.handle.net/10453/27110
dc.description.abstract	Metals often determine the chemical reactivity of the proteins to which they are bound. Each cell in the body tightly maintains a unique metalloproteomic profile, mostly dependent on function. This paper describes an analytical online flow injection quantitative size exclusion chromatography-inductively coupled plasma-mass spectrometry (SEC-ICP-MS) method, which was applied to profiling the metal-binding proteins found in primary cultures of neurons and astrocytes. This method can be conducted using similar amounts of sample to those used for Western blotting (20-150 μg protein), and has a turnaround time of <15 minutes. Metalloprotein standards for Fe (as ferritin), Cu and Zn (as superoxide dismutase-1) were used to construct multi-point calibration curves for online quantification of metalloproteins by SEC-ICP-MS. Homogenates of primary neuron and astrocyte cultures were analysed by SEC-ICP-MS. Online quantification by external calibration with metalloprotein standards determined the mass of metal eluting from the column relative to time (as pg s-1). Total on-column Fe, Cu and Zn detection limits ranged from 0.825 ± 0.005 ng to 13.6 ± 0.7 pg. Neurons and astrocytes exhibited distinct metalloprotein profiles, featuring both ubiquitous and unique metalloprotein species. Separation and detection by SEC-ICP-MS allows appraisal of these metalloproteins in their native state, and online quantification was achieved using this relatively simple external calibration process. © 2013 The Royal Society of Chemistry.	en_US
dc.relation.ispartof	Metallomics	en_US
dc.relation.isbasedon	10.1039/c3mt00227f	en_US
dc.subject.classification	Analytical Chemistry	en_US
dc.subject.mesh	Astrocytes	en_US
dc.subject.mesh	Neurons	en_US
dc.subject.mesh	Cells, Cultured	en_US
dc.subject.mesh	Animals	en_US
dc.subject.mesh	Mice	en_US
dc.subject.mesh	Copper	en_US
dc.subject.mesh	Iron	en_US
dc.subject.mesh	Zinc	en_US
dc.subject.mesh	Metalloproteins	en_US
dc.subject.mesh	Chromatography, Gel	en_US
dc.subject.mesh	Proteomics	en_US
dc.subject.mesh	Mass Spectrometry	en_US
dc.title	Profiling the iron, copper and zinc content in primary neuron and astrocyte cultures by rapid online quantitative size exclusion chromatography-inductively coupled plasma-mass spectrometry	en_US
dc.type	Journal Article
utslib.citation.volume	12	en_US
utslib.citation.volume	5	en_US
utslib.for	0301 Analytical Chemistry	en_US
utslib.for	03 Chemical Sciences	en_US
pubs.embargo.period	Not known	en_US
pubs.organisational-group	/University of Technology Sydney
pubs.organisational-group	/University of Technology Sydney/Faculty of Science
pubs.organisational-group	/University of Technology Sydney/Faculty of Science/School of Mathematical and Physical Sciences
pubs.organisational-group	/University of Technology Sydney/Strength - CFS - Centre for Forensic Science
utslib.copyright.status	open_access
pubs.issue	12	en_US
pubs.publication-status	Published	en_US
pubs.volume	5	en_US

Abstract:

Metals often determine the chemical reactivity of the proteins to which they are bound. Each cell in the body tightly maintains a unique metalloproteomic profile, mostly dependent on function. This paper describes an analytical online flow injection quantitative size exclusion chromatography-inductively coupled plasma-mass spectrometry (SEC-ICP-MS) method, which was applied to profiling the metal-binding proteins found in primary cultures of neurons and astrocytes. This method can be conducted using similar amounts of sample to those used for Western blotting (20-150 μg protein), and has a turnaround time of <15 minutes. Metalloprotein standards for Fe (as ferritin), Cu and Zn (as superoxide dismutase-1) were used to construct multi-point calibration curves for online quantification of metalloproteins by SEC-ICP-MS. Homogenates of primary neuron and astrocyte cultures were analysed by SEC-ICP-MS. Online quantification by external calibration with metalloprotein standards determined the mass of metal eluting from the column relative to time (as pg s-1). Total on-column Fe, Cu and Zn detection limits ranged from 0.825 ± 0.005 ng to 13.6 ± 0.7 pg. Neurons and astrocytes exhibited distinct metalloprotein profiles, featuring both ubiquitous and unique metalloprotein species. Separation and detection by SEC-ICP-MS allows appraisal of these metalloproteins in their native state, and online quantification was achieved using this relatively simple external calibration process. © 2013 The Royal Society of Chemistry.

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http://hdl.handle.net/10453/27110