TY - JOUR

T1 - Biomedical copper speciation in relation to Wilson's disease using strong anion exchange chromatography coupled to triple quadrupole inductively coupled plasma mass spectrometry

AU - Solovyev, Nikolay

AU - Ala, Aftab

AU - Schilsky, Michael

AU - Mills, Craig

AU - Willis, Karl

AU - Harrington, Chris F.

PY - 2020/2/15

Y1 - 2020/2/15

N2 - Biomedical analytical methods often rely on indirect measurements, such as immunoassays, which can lack effective metrological traceability. In the nephelometric determination of ceruloplasmin (Cp), an important protein whose circulating level is altered in Wilson's disease (WD), the anti-Cp antibody used is not specific for the biologically active holoprotein so the assay can overestimate the concentration of Cp due to the presence of the apoprotein. By providing quantitation using elemental standards, the use of strong anion exchange chromatography (SAX) coupled to triple quadrupole inductively coupled plasma mass spectrometry (ICP-MS-MS) can overcome the drawbacks of methods for the measurement of metalloproteins reliant on immunoassays. In the current study, a SAX-ICP-MS-MS method for Cp quantification was designed and tested in samples of blood serum of WD patients and healthy controls. Using standards based on a copper-EDTA complex for calibration, the method provides relatively simple quantification of Cp with the limit of detection of 0.1 μg L−1 (limit of quantification 0.4 μg L−1). The method was also used to investigate the copper species separated by using a 30 kDa cut-off ultrafiltration device. The so-called “exchangeable” copper fraction is considered as an alternative clinical biomarker of WD. Using the designed speciation approach, it was shown that the ultrafiltration method can overestimate the “exchangeable” copper fraction due to a removal of copper from Cp. This was confirmed by comparing the enzymatic activity of the fractions. Thus, the specificity of the “exchangeable” copper test can be ensured only under strict maintenance of ultrafiltration conditions.

AB - Biomedical analytical methods often rely on indirect measurements, such as immunoassays, which can lack effective metrological traceability. In the nephelometric determination of ceruloplasmin (Cp), an important protein whose circulating level is altered in Wilson's disease (WD), the anti-Cp antibody used is not specific for the biologically active holoprotein so the assay can overestimate the concentration of Cp due to the presence of the apoprotein. By providing quantitation using elemental standards, the use of strong anion exchange chromatography (SAX) coupled to triple quadrupole inductively coupled plasma mass spectrometry (ICP-MS-MS) can overcome the drawbacks of methods for the measurement of metalloproteins reliant on immunoassays. In the current study, a SAX-ICP-MS-MS method for Cp quantification was designed and tested in samples of blood serum of WD patients and healthy controls. Using standards based on a copper-EDTA complex for calibration, the method provides relatively simple quantification of Cp with the limit of detection of 0.1 μg L−1 (limit of quantification 0.4 μg L−1). The method was also used to investigate the copper species separated by using a 30 kDa cut-off ultrafiltration device. The so-called “exchangeable” copper fraction is considered as an alternative clinical biomarker of WD. Using the designed speciation approach, it was shown that the ultrafiltration method can overestimate the “exchangeable” copper fraction due to a removal of copper from Cp. This was confirmed by comparing the enzymatic activity of the fractions. Thus, the specificity of the “exchangeable” copper test can be ensured only under strict maintenance of ultrafiltration conditions.

KW - Blood serum

KW - Ceruloplasmin

KW - Copper

KW - Inductively coupled plasma mass spectrometry

KW - Strong anion exchange chromatography

KW - Wilson's disease

KW - Hepatolenticular Degeneration/blood

KW - Copper/blood

KW - Humans

KW - Biomedical Research

KW - Chromatography, Ion Exchange/instrumentation

KW - Mass Spectrometry/instrumentation

KW - FRACTIONATION

KW - METALLOPROTEINS

KW - CERULOPLASMIN

KW - GUIDELINES

KW - SERUM

KW - QUANTITATIVE-ANALYSIS

KW - ICP-MS

KW - ABSOLUTE QUANTIFICATION

KW - PROTEINS

KW - TOOL

UR - http://www.scopus.com/inward/record.url?scp=85075902275&partnerID=8YFLogxK

U2 - 10.1016/j.aca.2019.11.033

DO - 10.1016/j.aca.2019.11.033

M3 - Article

C2 - 31948584

AN - SCOPUS:85075902275

VL - 1098

SP - 27

EP - 36

JO - Analytica Chimica Acta

JF - Analytica Chimica Acta

SN - 0003-2670

ER -