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Article: Accurate characterization of β-amyloid (Aβ40, Aβ42) standards using species-specific isotope dilution by means of HPLC-ICP-MS/MS

Schaier, Martin / Hermann, Gerrit / Koellensperger, Gunda / Theiner, Sarah

Analytical and bioanalytical chemistry. 2022 Jan., v. 414, no. 1

2022

Abstract: The amyloid β peptide, as one of the main components in senile plaque, represents a defining pathological feature for Alzheimer’s disease, and is therefore commonly used as a biomarker for this disease in clinical analysis. However, the selection of ... ...

Abstract	The amyloid β peptide, as one of the main components in senile plaque, represents a defining pathological feature for Alzheimer’s disease, and is therefore commonly used as a biomarker for this disease in clinical analysis. However, the selection of suitable standards is limited here, since only a few are commercially available, and these suffer from varying purity. Hence, the accurate characterization of these standards is of great importance. In this study, we developed a method for the traceable quantification of the peptide content using species-specific isotope dilution and ICP-MS/MS detection. It is based on the separation of the sulfur-containing amino acids methionine and cysteine after oxidation and hydrolysis of the peptide. Using a strong anion exchange column, both amino acids could be separated from each other, as well as from their oxidized forms and sulfate. The sulfur content was determined via ICP-MS/MS using oxygen as reaction gas. Species-specific isotope dilution was enabled by using a ³⁴S-labeled yeast hydrolysate, containing methionine sulfone and cysteic acid with different isotopic composition. The peptide contents of Aβ standards (Aβ40,42), as well as myoglobin and lysozyme with different degrees of purity, were determined. For validation purposes, the standard reference material NIST 2389a, which contains the amino acids in a similar concentration, was subjected to the developed sample preparation and analysis method. In addition to accounting for errors during sample preparation, high levels of accuracy and precision could be obtained using this method, making it fit-for-purpose for the characterization of peptide standards.
Keywords	amyloid ; analytical chemistry ; anion exchange ; biomarkers ; cysteic acid ; cysteine ; hydrolysates ; hydrolysis ; isotope dilution technique ; lysozyme ; methionine ; myoglobin ; oxidation ; oxygen ; peptides ; sulfates ; sulfur ; yeasts
Language	English
Dates of publication	2022-01
Size	p. 639-648.
Publishing place	Springer Berlin Heidelberg
Document type	Article
ISSN	1618-2642
DOI	10.1007/s00216-021-03571-6
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: Multiparametric Tissue Characterization Utilizing the Cellular Metallome and Immuno-Mass Spectrometry Imaging.

Schaier, Martin / Theiner, Sarah / Baier, Dina / Braun, Gabriel / Berger, Walter / Koellensperger, Gunda

JACS Au

2023 Volume 3, Issue 2, Page(s) 419–428

Abstract: In this study, we present a workflow that enables spatial single-cell metallomics in tissue decoding the cellular heterogeneity. Low-dispersion laser ablation in combination with inductively coupled plasma time-of-flight mass spectrometry (LA-ICP-TOFMS) ... ...

Abstract	In this study, we present a workflow that enables spatial single-cell metallomics in tissue decoding the cellular heterogeneity. Low-dispersion laser ablation in combination with inductively coupled plasma time-of-flight mass spectrometry (LA-ICP-TOFMS) provides mapping of endogenous elements with cellular resolution at unprecedented speed. Capturing the heterogeneity of the cellular population by metals only is of limited use as the cell type, functionality, and cell state remain elusive. Therefore, we expanded the toolbox of single-cell metallomics by integrating the concepts of imaging mass cytometry (IMC). This multiparametric assay successfully utilizes metal-labeled antibodies for cellular tissue profiling. One important challenge is the need to preserve the original metallome in the sample upon immunostaining. Therefore, we studied the impact of extensive labeling on the obtained endogenous cellular ionome data by quantifying elemental levels in consecutive tissue sections (with and without immunostaining) and correlating elements with structural markers and histological features. Our experiments showed that the elemental tissue distribution remained intact for selected elements such as sodium, phosphorus, and iron, while absolute quantification was precluded. We hypothesize that this integrated assay not only advances single-cell metallomics (enabling to link metal accumulation to multi-dimensional characterization of cells/cell populations), but in turn also enhances selectivity in IMC, as in selected cases, labeling strategies can be validated by elemental data. We showcase the power of this integrated single-cell toolbox using an in vivo tumor model in mice and provide mapping of the sodium and iron homeostasis as linked to different cell types and function in mouse organs (such as spleen, kidney, and liver). Phosphorus distribution maps added structural information, paralleled by the DNA intercalator visualizing the cellular nuclei. Overall, iron imaging was the most relevant addition to IMC. In tumor samples, for example, iron-rich regions correlated with high proliferation and/or located blood vessels, which are key for potential drug delivery.
Language	English
Publishing date	2023-02-08
Publishing country	United States
Document type	Journal Article
ISSN	2691-3704
ISSN (online)	2691-3704
DOI	10.1021/jacsau.2c00571
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Accurate characterization of β-amyloid (Aβ40, Aβ42) standards using species-specific isotope dilution by means of HPLC-ICP-MS/MS.

Schaier, Martin / Hermann, Gerrit / Koellensperger, Gunda / Theiner, Sarah

Analytical and bioanalytical chemistry

2021 Volume 414, Issue 1, Page(s) 639–648

Abstract: The amyloid β peptide, as one of the main components in senile plaque, represents a defining pathological feature for Alzheimer's disease, and is therefore commonly used as a biomarker for this disease in clinical analysis. However, the selection of ... ...

Abstract	The amyloid β peptide, as one of the main components in senile plaque, represents a defining pathological feature for Alzheimer's disease, and is therefore commonly used as a biomarker for this disease in clinical analysis. However, the selection of suitable standards is limited here, since only a few are commercially available, and these suffer from varying purity. Hence, the accurate characterization of these standards is of great importance. In this study, we developed a method for the traceable quantification of the peptide content using species-specific isotope dilution and ICP-MS/MS detection. It is based on the separation of the sulfur-containing amino acids methionine and cysteine after oxidation and hydrolysis of the peptide. Using a strong anion exchange column, both amino acids could be separated from each other, as well as from their oxidized forms and sulfate. The sulfur content was determined via ICP-MS/MS using oxygen as reaction gas. Species-specific isotope dilution was enabled by using a
MeSH term(s)	Amyloid beta-Peptides ; Chromatography, High Pressure Liquid ; Isotopes ; Peptide Fragments ; Tandem Mass Spectrometry/methods
Chemical Substances	Amyloid beta-Peptides ; Isotopes ; Peptide Fragments ; amyloid beta-protein (1-42)
Language	English
Publishing date	2021-08-06
Publishing country	Germany
Document type	Journal Article
ZDB-ID	201093-8
ISSN	1618-2650 ; 0016-1152 ; 0372-7920
ISSN (online)	1618-2650
ISSN	0016-1152 ; 0372-7920
DOI	10.1007/s00216-021-03571-6
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Article ; Online: Semiquantitative Analysis for High-Speed Mapping Applications of Biological Samples Using LA-ICP-TOFMS

Metarapi, Dino / Schweikert, Andreas / Jerše, Ana / Schaier, Martin / van Elteren, Johannes T. / Koellensperger, Gunda / Theiner, Sarah / Šala, Martin

Analytical Chemistry. 2023 Apr. 26, v. 95, no. 19 p.7804-7812

2023

Abstract: Laser ablation (LA) in combination with inductively coupled plasma time-of-flight mass spectrometry (ICP-TOFMS) enables monitoring of elements from the entire mass range for every pixel, regardless of the isotopes of interest for a certain application. ... ...

Abstract	Laser ablation (LA) in combination with inductively coupled plasma time-of-flight mass spectrometry (ICP-TOFMS) enables monitoring of elements from the entire mass range for every pixel, regardless of the isotopes of interest for a certain application. This provides nontargeted multi-element (bio-)imaging capabilities and the unique possibility to screen for elements that were initially not expected in the sample. Quantification of a large range of elements is limited as the preparation of highly multiplexed calibration standards for bioimaging applications by LA-ICP-(TOF)MS is challenging. In this study, we have developed a workflow for semiquantitative analysis by LA-ICP-TOFMS based on multi-element gelatin micro-droplet standards. The presented approach is intended for the mapping of biological samples due to the requirement of matrix-matched standards for accurate quantification in LA-ICPMS, a prerequisite that is given by the use of gelatin-based standards. A library of response factors was constructed based on 72 elements for the semiquantitative calculations. The presented method was evaluated in two stages: (i) on gelatin samples with known elemental concentrations and (ii) on real-world samples that included prime examples of bioimaging (mouse spleen and tumor tissue). The developed semiquantification approach was based on 10 elements as calibration standards and provided the determination of 136 nuclides of 63 elements, with errors below 25%, and for half of the nuclides, below 10%. A web application for quantification and semiquantification of LA-ICP(-TOF)MS data was developed, and a detailed description is presented to easily allow others to use the presented method.
Keywords	Internet ; analytical chemistry ; bioimaging ; calibration ; gelatin ; mass spectrometry ; mice ; neoplasms ; spleen
Language	English
Dates of publication	2023-0426
Size	p. 7804-7812.
Publishing place	American Chemical Society
Document type	Article ; Online
ZDB-ID	1508-8
ISSN	1520-6882 ; 0003-2700
ISSN (online)	1520-6882
ISSN	0003-2700
DOI	10.1021/acs.analchem.3c01439
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: Semiquantitative Analysis for High-Speed Mapping Applications of Biological Samples Using LA-ICP-TOFMS.

Metarapi, Dino / Schweikert, Andreas / Jerše, Ana / Schaier, Martin / van Elteren, Johannes T / Koellensperger, Gunda / Theiner, Sarah / Šala, Martin

Analytical chemistry

2023 Volume 95, Issue 19, Page(s) 7804–7812

Abstract	Laser ablation (LA) in combination with inductively coupled plasma time-of-flight mass spectrometry (ICP-TOFMS) enables monitoring of elements from the entire mass range for every pixel, regardless of the isotopes of interest for a certain application. This provides nontargeted multi-element (bio-)imaging capabilities and the unique possibility to screen for elements that were initially not expected in the sample. Quantification of a large range of elements is limited as the preparation of highly multiplexed calibration standards for bioimaging applications by LA-ICP-(TOF)MS is challenging. In this study, we have developed a workflow for semiquantitative analysis by LA-ICP-TOFMS based on multi-element gelatin micro-droplet standards. The presented approach is intended for the mapping of biological samples due to the requirement of matrix-matched standards for accurate quantification in LA-ICPMS, a prerequisite that is given by the use of gelatin-based standards. A library of response factors was constructed based on 72 elements for the semiquantitative calculations. The presented method was evaluated in two stages: (i) on gelatin samples with known elemental concentrations and (ii) on real-world samples that included prime examples of bioimaging (mouse spleen and tumor tissue). The developed semiquantification approach was based on 10 elements as calibration standards and provided the determination of 136 nuclides of 63 elements, with errors below 25%, and for half of the nuclides, below 10%. A web application for quantification and semiquantification of LA-ICP(-TOF)MS data was developed, and a detailed description is presented to easily allow others to use the presented method.
MeSH term(s)	Mice ; Animals ; Mass Spectrometry/methods ; Gelatin ; Laser Therapy ; Spectrum Analysis ; Food
Chemical Substances	Gelatin (9000-70-8)
Language	English
Publishing date	2023-04-25
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1508-8
ISSN	1520-6882 ; 0003-2700
ISSN (online)	1520-6882
ISSN	0003-2700
DOI	10.1021/acs.analchem.3c01439
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article: Elemental Mapping of Human Malignant Mesothelioma Tissue Samples Using High-Speed LA–ICP–TOFMS Imaging

Voloaca, Oana M. / Clench, Malcolm R. / Koellensperger, Gunda / Cole, Laura M. / Haywood-Small, Sarah L. / Theiner, Sarah

Analytical chemistry. 2022 Jan. 24, v. 94, no. 5

2022

Abstract: This is the first report of the use of laser ablation–inductively coupled plasma time-of-flight mass spectrometry (LA–ICP–TOFMS) to analyze human malignant pleural mesothelioma (MPM) samples at the cellular level. MPM is an aggressive, incurable cancer ... ...

Abstract	This is the first report of the use of laser ablation–inductively coupled plasma time-of-flight mass spectrometry (LA–ICP–TOFMS) to analyze human malignant pleural mesothelioma (MPM) samples at the cellular level. MPM is an aggressive, incurable cancer associated with asbestos exposure, with a long latency and poor overall survival. Following careful optimization of the laser fluence, the simultaneous ablation of soft biological tissue and hard mineral fibers was possible, allowing the spatial detection of elements such as Si, Mg, Ca, and Fe, which are also present in the glass substrate. A low-dispersion LA setup was employed, which provided the high spatial resolution necessary to identify the asbestos fibers and fiber fragments in the tissue and to characterize the metallome at the cellular level (a pixel size of 2 μm), with a high speed (at 250 Hz). The multielement LA–ICP–TOFMS imaging approach enabled (i) the detection of asbestos fibers/mineral impurities within the MPM tissue samples of patients, (ii) the visualization of the tissue structure with the endogenous elemental pattern at high spatial resolution, and (iii) obtaining insights into the metallome of MPM patients with different pathologies in a single analysis run. Asbestos and other mineral fibers were detected in the lung and pleura tissue of MPM patients, respectively, based on their multielement pattern (Si, Mg, Ca, Fe, and Sr). Interestingly, strontium was detected in asbestos fibers, suggesting a link between this potential toxic element and MPM pathogenesis. Furthermore, monitoring the metallome around the talc deposit regions (characterized by elevated levels of Al, Mg, and Si) revealed significant tissue damage and inflammation caused by talc pleurodesis. LA–ICP–TOFMS results correlated to Perls’ Prussian blue and histological staining of the corresponding serial sections. Ultimately, the ultra-high-speed and high-spatial-resolution capabilities of this novel LA–ICP–TOFMS setup may become an important clinical tool for simultaneous asbestos detection, metallome monitoring, and biomarker identification.
Keywords	analytical chemistry ; asbestos ; biomarkers ; glass ; histology ; humans ; inflammation ; lungs ; mass spectrometry ; mesothelioma ; metallome ; pathogenesis ; pleura ; strontium ; talc ; toxicity
Language	English
Dates of publication	2022-0124
Size	p. 2597-2606.
Publishing place	American Chemical Society
Document type	Article
ZDB-ID	1508-8
ISSN	1520-6882 ; 0003-2700
ISSN (online)	1520-6882
ISSN	0003-2700
DOI	10.1021/acs.analchem.1c04857
Database	NAL-Catalogue (AGRICOLA)

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Article: Micro-droplet-based calibration for quantitative elemental bioimaging by LA-ICPMS

Schweikert, Andreas / Theiner, Sarah / Wernitznig, Debora / Schoeberl, Anna / Schaier, Martin / Neumayer, Sophie / Keppler, Bernhard K. / Koellensperger, Gunda

Analytical and bioanalytical chemistry. 2022 Jan., v. 414, no. 1

2022

Abstract: In this work, a novel standardization strategy for quantitative elemental bioimaging is evaluated. More specifically, multi-element quantification by laser ablation-inductively coupled plasma-time-of-flight mass spectrometry (LA-ICP-TOFMS) is performed ... ...

Abstract	In this work, a novel standardization strategy for quantitative elemental bioimaging is evaluated. More specifically, multi-element quantification by laser ablation-inductively coupled plasma-time-of-flight mass spectrometry (LA-ICP-TOFMS) is performed by multi-point calibration using gelatin-based micro-droplet standards and validated using in-house produced reference materials. Fully automated deposition of micro-droplets by micro-spotting ensured precise standard volumes of 400 ± 5 pL resulting in droplet sizes of around 200 μm in diameter. The small dimensions of the micro-droplet standards and the use of a low-dispersion laser ablation setup reduced the analysis time required for calibration by LA-ICPMS significantly. Therefore, as a key advance, high-throughput analysis (pixel acquisition rates of more than 200 Hz) enabled to establish imaging measurement sequences with quality control- and standardization samples comparable to solution-based quantification exercises by ICP-MS. Analytical figures of merit such as limit of detection, precision, and accuracy of the calibration approach were assessed for platinum and for elements with biological key functions from the lower mass range (phosphorus, copper, and zinc). As a proof-of-concept application, the tool-set was employed to investigate the accumulation of metal-based anticancer drugs in multicellular tumor spheroid models at clinically relevant concentrations. Graphical abstract
Keywords	analytical chemistry ; bioimaging ; calibration ; copper ; detection limit ; droplets ; mass spectrometry ; neoplasms ; phosphorus ; platinum ; zinc
Language	English
Dates of publication	2022-01
Size	p. 485-495.
Publishing place	Springer Berlin Heidelberg
Document type	Article
ISSN	1618-2642
DOI	10.1007/s00216-021-03357-w
Database	NAL-Catalogue (AGRICOLA)

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Article: Cisplatin Uptake in Macrophage Subtypes at the Single-Cell Level by LA-ICP-TOFMS Imaging

Schoeberl, Anna / Gutmann, Michael / Theiner, Sarah / Schaier, Martin / Schweikert, Andreas / Berger, Walter / Koellensperger, Gunda

Analytical chemistry. 2021 Nov. 30, v. 93, no. 49

2021

Abstract: A high-throughput laser ablation–inductively coupled plasma–time-of-flight mass spectrometry (LA-ICP-TOFMS) workflow was implemented for quantitative single-cell analysis following cytospin preparation of cells. For the first time, in vitro studies on ... ...

Abstract	A high-throughput laser ablation–inductively coupled plasma–time-of-flight mass spectrometry (LA-ICP-TOFMS) workflow was implemented for quantitative single-cell analysis following cytospin preparation of cells. For the first time, in vitro studies on cisplatin exposure addressed human monocytes and monocyte-derived macrophages (undifferentiated THP-1 monocytic cells, differentiated M0 macrophages, as well as further polarized M1 and M2 phenotypes) at the single-cell level. The models are of particular interest as macrophages comprise the biggest part of immune cells present in the tumor microenvironment and play an important role in modulating tumor growth and progression. The introduced bioimaging workflow proved to be universally applicable to adherent and suspension cell cultures and fit-for-purpose for the quantitative analysis of several hundreds of cells within minutes. Both, cross-validation of the method with single-cell analysis in suspension for THP-1 cells and with LA-ICP-TOFMS analysis of adherent M0 cells grown on chambered glass coverslips, revealed agreeing platinum concentrations at the single-cell level. A high incorporation of cisplatin was observed in M2 macrophages compared to the M0 and M1 macrophage subtypes and the monocyte model, THP-1. The combination with bright-field images and monitoring of highly abundant endogenous elements such as phosphorus and sodium at a high spatial resolution allowed assessing cell size and important morphological cell parameters and thus straightforward control over several cell conditions. This way, apoptotic cells and cell debris as well as doublets or cell clusters could be easily excluded prior to data evaluation without additional staining.
Keywords	analytical chemistry ; apoptosis ; bioimaging ; cisplatin ; glass ; humans ; macrophages ; mass spectrometry ; models ; monocytes ; neoplasms ; phosphorus ; platinum ; quantitative analysis ; sodium
Language	English
Dates of publication	2021-1130
Size	p. 16456-16465.
Publishing place	American Chemical Society
Document type	Article
ZDB-ID	1508-8
ISSN	1520-6882 ; 0003-2700
ISSN (online)	1520-6882
ISSN	0003-2700
DOI	10.1021/acs.analchem.1c03442
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Article ; Online: Laser ablation-ICP-TOFMS imaging of germ cell tumors of patients undergoing platinum-based chemotherapy.

Theiner, Sarah / Schweikert, Andreas / Haberler, Christine / Peyrl, Andreas / Koellensperger, Gunda

Metallomics : integrated biometal science

2020 Volume 12, Issue 8, Page(s) 1246–1252

Abstract: A low dispersion laser ablation setup in combination with inductively coupled plasma-time-of-flight mass spectrometry (LA-ICP-TOFMS) was applied to clinical samples of patients undergoing platinum-based chemotherapy. The platinum accumulation together ... ...

Abstract	A low dispersion laser ablation setup in combination with inductively coupled plasma-time-of-flight mass spectrometry (LA-ICP-TOFMS) was applied to clinical samples of patients undergoing platinum-based chemotherapy. The platinum accumulation together with the distribution of elements with biological key functions (Mg, P, S, Ca, Fe, Cu and Zn) was studied in central nervous system germ cell tumor (CNS GCT) tissue, which is an aggressive tumor type located in the brain. Heterogeneous elemental distribution patterns were obtained with a pixel size of 10 μm and were correlated to histological analysis of serial sections using hematoxylin eosin staining. Highest platinum accumulation correlated with areas of necrosis, which exhibited high levels of magnesium, sulphur and calcium. Small traces of gadolinium were found in the tumor sections, which is a result of prior magnetic resonance imaging. Iron accumulated in regions, which were dense in blood vessels, whereas areas with fibrosis scar showed the lowest levels of all detected elements. This LA-ICP-TOFMS study demonstrates that the chemotherapeutic drug cisplatin accumulated in the germ cell tumor located in the brain, which is also reflected by the therapy response of the patients.
MeSH term(s)	Calcium/chemistry ; Laser Therapy ; Magnesium/chemistry ; Mass Spectrometry ; Platinum/chemistry
Chemical Substances	Platinum (49DFR088MY) ; Magnesium (I38ZP9992A) ; Calcium (SY7Q814VUP)
Language	English
Publishing date	2020-06-07
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2474317-3
ISSN	1756-591X ; 1756-5901
ISSN (online)	1756-591X
ISSN	1756-5901
DOI	10.1039/d0mt00080a
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Cisplatin Uptake in Macrophage Subtypes at the Single-Cell Level by LA-ICP-TOFMS Imaging.

Schoeberl, Anna / Gutmann, Michael / Theiner, Sarah / Schaier, Martin / Schweikert, Andreas / Berger, Walter / Koellensperger, Gunda

Analytical chemistry

2021 Volume 93, Issue 49, Page(s) 16456–16465

Abstract: A high-throughput laser ablation-inductively coupled plasma-time-of-flight mass spectrometry (LA-ICP-TOFMS) workflow was implemented for quantitative single-cell analysis following cytospin preparation of cells. For the first time, in vitro studies on ... ...

Abstract	A high-throughput laser ablation-inductively coupled plasma-time-of-flight mass spectrometry (LA-ICP-TOFMS) workflow was implemented for quantitative single-cell analysis following cytospin preparation of cells. For the first time, in vitro studies on cisplatin exposure addressed human monocytes and monocyte-derived macrophages (undifferentiated THP-1 monocytic cells, differentiated M0 macrophages, as well as further polarized M1 and M2 phenotypes) at the single-cell level. The models are of particular interest as macrophages comprise the biggest part of immune cells present in the tumor microenvironment and play an important role in modulating tumor growth and progression. The introduced bioimaging workflow proved to be universally applicable to adherent and suspension cell cultures and fit-for-purpose for the quantitative analysis of several hundreds of cells within minutes. Both, cross-validation of the method with single-cell analysis in suspension for THP-1 cells and with LA-ICP-TOFMS analysis of adherent M0 cells grown on chambered glass coverslips, revealed agreeing platinum concentrations at the single-cell level. A high incorporation of cisplatin was observed in M2 macrophages compared to the M0 and M1 macrophage subtypes and the monocyte model, THP-1. The combination with bright-field images and monitoring of highly abundant endogenous elements such as phosphorus and sodium at a high spatial resolution allowed assessing cell size and important morphological cell parameters and thus straightforward control over several cell conditions. This way, apoptotic cells and cell debris as well as doublets or cell clusters could be easily excluded prior to data evaluation without additional staining.
MeSH term(s)	Cisplatin/pharmacology ; Humans ; Macrophages ; Monocytes ; Neuroblastoma ; THP-1 Cells ; Tumor Microenvironment
Chemical Substances	Cisplatin (Q20Q21Q62J)
Language	English
Publishing date	2021-11-30
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1508-8
ISSN	1520-6882 ; 0003-2700
ISSN (online)	1520-6882
ISSN	0003-2700
DOI	10.1021/acs.analchem.1c03442
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