Mass spectrometry or immunoassay: est modus in rebus

Antonelli, Giorgia; Marinova, Mariela; Artusi, Carlo Alberto; Plebani, Mario

doi:10.1515/cclm-2017-0197

Cited by 19 publications

(12 citation statements)

References 28 publications

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“…However, immunoassays do not exist for all clinically relevant biomarkers. These assays often reported a lack of reproducibility across different platforms and sometimes a lack of specificity due to the antibodies' cross-reactivity [3]. Liquid chromatography coupled to mass spectrometry is a powerful alternative analytical technique in bioanalysis because of its good reproducibility and high specificity by eliminating interferences that may impact immunoassays (e.g., cross-reactivity of antibodies) [3].…”

Section: Introductionmentioning

confidence: 99%

“…These assays often reported a lack of reproducibility across different platforms and sometimes a lack of specificity due to the antibodies' cross-reactivity [3]. Liquid chromatography coupled to mass spectrometry is a powerful alternative analytical technique in bioanalysis because of its good reproducibility and high specificity by eliminating interferences that may impact immunoassays (e.g., cross-reactivity of antibodies) [3]. However, LC-MS requires a high initial investment for instrument acquisition and highly experienced operators, and, for intended use in clinical practice, presents a limited sample throughput compared to immunoassays [3][4][5].…”

Section: Introductionmentioning

confidence: 99%

“…Liquid chromatography coupled to mass spectrometry is a powerful alternative analytical technique in bioanalysis because of its good reproducibility and high specificity by eliminating interferences that may impact immunoassays (e.g., cross-reactivity of antibodies) [3]. However, LC-MS requires a high initial investment for instrument acquisition and highly experienced operators, and, for intended use in clinical practice, presents a limited sample throughput compared to immunoassays [3][4][5]. However, this last point tends to be addressed by developing high-throughput LC-MS methods, particularly on sample preparation steps, to improve LC-MS adoption into routine practice.…”

Section: Introductionmentioning

confidence: 99%

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Development of an antibody-free ID-LC MS method for the quantification of procalcitonin in human serum at sub-microgram per liter level using a peptide-based calibration

et al. 2021

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The quantification of low abundant proteins in complex matrices by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) remains challenging. A measurement procedure based on optimized antibody-free sample preparation and isotope dilution coupled to LC-MS/MS was developed to quantify procalcitonin (PCT) in human serum at sub-microgram per liter level. A combination of sodium deoxycholate-assisted protein precipitation with acetonitrile, solid-phase extraction, and trypsin digestion assisted with Tween-20 enhanced the method sensitivity. Linearity was established through peptide-based calibration curves in the serum matrix (0.092-5.222 μg/L of PCT) with a good linear fit (R2 ≥ 0.999). Quality control materials spiked with known amounts of protein-based standards were used to evaluate the method's accuracy. The bias ranged from −2.6 to +4.3%, and the intra-day and inter-day coefficients of variations (CVs) were below 2.2% for peptide-based quality controls. A well-characterized correction factor was determined and applied to compensate for digestion incompleteness and material loss before the internal standards spike. Results with metrological traceability to the SI units were established using standard peptide of well-characterized purity determined by peptide impurity corrected amino acid analysis. The validated method enables accurate quantification of PCT in human serum at a limit of quantification down to 0.245 μg/L (bias −1.9%, precision 9.1%). The method was successfully applied to serum samples obtained from patients with sepsis. Interestingly, the PCT concentration reported implementing the isotope dilution LC-MS/MS method was twofold lower than the concentration provided by an immunoassay.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Development of an antibody-free ID-LC MS method for the quantification of procalcitonin in human serum at sub-microgram per liter level using a peptide-based calibration

et al. 2021

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“…However, they present important limitations when used in the initial study of new biomarkers, especially due to their relatively low specificity, cross-reactivity and lack of agreement between platforms. 26 LC-MS/MS emerges as the methodology of choice to evaluate the potential of a new biomarker, such as free 25OHD, that depends directly on the determination of DBP concentrations. Therefore, we developed a method for the phenotyping and relative quantification of DBP by LC-MS/MS, which is similar to the one published by Henderson et al 20 However, we decided to focus on paediatric population to elucidate if there are differences in the DBP concentrations among phenotypes.…”

Section: Discussionmentioning

confidence: 99%

“…19 To contribute to clarifying these facts, the aim of this study was to develop a mass spectrometry-based method for phenotype identification and relative quantification of DBP, in order to avoid many of the problems associated with immunoassays. 26…”

Section: Introductionmentioning

confidence: 99%

Phenotyping and relative quantification of vitamin D binding protein in a paediatric population by using liquid chromatography–tandem mass spectrometry

et al. 2018

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Background Adequate concentrations of vitamin D are required to ensure bone health and minimize the incidence of multiple extraskeletal diseases. Although total 25-hydroxyvitamin D (25OHD) remains the recommended biomarker for assessing vitamin D status, it has been speculated that free 25OHD correlates better with clinical outcomes. The calculation of free 25OHD depends on the concentrations of vitamin D binding protein (DBP), the determination of which involves different immunoassays and has led to varying results and conclusions. We developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous identification and relative quantification of DBP isoforms. Methods We used serum samples from healthy children ( n = 79), mainly Caucasian (88%). Proteins were denatured, reduced, alkylated and digested with trypsin. Purified peptides were analysed by LC-MS/MS. The DBP phenotype was established by using the combinations of tryptic peptides associated with each of the three isoforms and one peptide common to all of them to perform relative quantification. The genotyping of volunteers ( n = 7) facilitated verification of the ability of our method to correctly identify the DBP phenotype. Results The DBP phenotype was correctly established in all samples from volunteers, based on the 100% correlation observed with the genotype. The most common DBP phenotype in Caucasian children was 2/1S (34%) and the rarest 1F/1F (2%). The relative quantification of DBP concentrations did not show statistically significant differences between phenotypes ( P = 0.11). Conclusions LC-MS/MS enabled simultaneous phenotyping and relative quantification of DBP, while avoiding the analytical limitations of immunoassays and confirming similar concentrations of DBP in all phenotypes.

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Measurement of urinary free cortisol by LC–MS–MS: adoption of a literature reference range and comparison with our current immunometric method

Bianchi

Campi

Sessa

et al. 2019

J Endocrinol Invest

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Mass spectrometry or immunoassay: est modus in rebus

Cited by 19 publications

References 28 publications

Development of an antibody-free ID-LC MS method for the quantification of procalcitonin in human serum at sub-microgram per liter level using a peptide-based calibration

Development of an antibody-free ID-LC MS method for the quantification of procalcitonin in human serum at sub-microgram per liter level using a peptide-based calibration

Phenotyping and relative quantification of vitamin D binding protein in a paediatric population by using liquid chromatography–tandem mass spectrometry

Measurement of urinary free cortisol by LC–MS–MS: adoption of a literature reference range and comparison with our current immunometric method

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