Multiplexed Immunoassay: Quantitation and Profiling of Serum Biomarkers Using Magnetic Nanoprobes and MALDI-TOF MS

Wang, Kai Yi; Chuang, Szu An; Lin, Po Hsun; Huang, Li-Shing; Chen, Shu Hua; Ouarda, Saib; Pan, Wen Harn; Lee, Ping Ying; Lin, Chun Cheng; Chen, Yu‐Ju

doi:10.1021/ac800354u

Cited by 77 publications

(61 citation statements)

References 45 publications

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“…Efforts at quantification using MALDI-MS 32,33 have been published but are inherently encumbered by the higher-order quantitative limitations. Nonetheless, these methods are often multiplexed and offer rapid, alternative approaches for the generation of lowerorder methods for more routine analyses.…”

Section: Discussionmentioning

confidence: 99%

Reference Measurement Procedure Development for C-Reactive Protein in Human Serum

Kilpatrick

Bunk

2009

Anal. Chem.

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This paper describes the development of a reference measurement procedure to quantify human C-reactive protein (CRP) in serum using affinity techniques prior to tryptic digestion and liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the certification of reference materials in clinically relevant ranges. The absence of a suitable internal standard for the CRP measurement, necessary to eliminate potential measurement bias in both the affinity purification and trypsin digestion steps, was addressed using the method of standard addition. The standard addition quantification approach was combined with affinity purification, using an anti-CRP monoclonal antibody conjugated to polystyrene beads, trypsin digestion of the purified protein, and LC-MS/MS analysis of CRP tryptic peptides. The effectiveness of intact protein affinity purification was evaluated through the measurement of CRP in several serum-based CRP control materials, yielding levels that were comparable to their expected mean concentration values. Quantitative results were confirmed with an external calibration approach. This study demonstrates the feasibility of affinity purification with LC-MS/MS for the reference measurement procedure development of low abundance serum protein analytes.Reference measurement procedures are used for a variety of purposes including (1) the assessment of the measurement quality of routine measurement procedures, (2) the value assignment of high-order calibration solutions (the so-called "master calibrators") for routine assays, and (3) the value assignment of certified reference materials. 1,2 In these roles, reference measurement procedures are considered "higher-order" measurement procedures, as they aim to achieve a higher level of accuracy and precision than the routine assay for the analyte being measured, producing a value with a low uncertainty of defined magnitude. In order for a reference measurement procedure to be fit for this purpose, it must be thoroughly evaluated to identify sources of bias and assess their magnitude. [3][4][5] Minimizing or eliminating bias is necessary to measure the most accurate, true concentration of the analyte. Because of the need for high levels of accuracy and precision, reference measurement procedures are typically low-throughput, labor intensive, and often costly to perform, in sharp contrast to what is desirable for routine measurements.Reference measurement procedures are key elements in the establishment of measurement standardization and metrological traceability in clinical chemistry. For more than 30 years, isotope dilution mass spectrometry (IDMS) has been one of the analytical techniques used frequently in reference measurement procedures, particularly for small molecule and inorganic analytes of clinical importance. 6 Despite this, few reference measurement procedures exist for clinically relevant proteins. This is one reason why so many clinical protein analytes are measured in arbitrary International Units (IUs); without a reference measurement procedure to...

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Section: Discussionmentioning

confidence: 99%

Reference Measurement Procedure Development for C-Reactive Protein in Human Serum

Kilpatrick

Bunk

2009

Anal. Chem.

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“…Antigen-antibody complexes in the 150-400 kDa range were chemically cross-linked and directly detected by MALDI mass spectrometry (Nazabal, Wenzel, & Zenobi, 2006). Affinity chromatography can provide biomarkers, therapeutic targets or therapeutic agents in a pure enough state that its tryptic fragments can be characterized and quantified (Chou et al, 2005;Christoffersen et al, 2006;Wang et al, 2006Wang et al, , 2008bZhao et al, 2006;Kiernan et al, 2006a;Kiernan, Nedelkov, & Nelson, 2006b;Cole et al, 2007;Csanky et al, 2007;Gatlin-Bunai et al, 2007;Heo et al, 2007;Keshishian et al, 2007;Saito & Munakata, 2007;Whiteaker et al, 2007c;Berna et al, 2008;Borges et al, 2008;Dubois et al, 2008;Calvano, Zambonin, & Jensen, 2008;Cederfur et al, 2008;Kulasingam et al, 2008). Natriuretic peptides and the receptor isoforms (Rubattu et al, 2006(Rubattu et al, , 2007 were isolated by classic purifications (Baines, DeBold, & Sonnenberg, 1983) and the pro-B-type natriuretic peptides can be captured by antibodies and detected with mass spectrometry (Hammerer-Lercher et al, 2008).…”

Section: Affinity Chromatographymentioning

confidence: 99%

Mass spectrometry of peptides and proteins from human blood

Zhu

Bowden

Zhang

et al. 2010

Mass Spectrometry Reviews

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It is difficult to convey the accelerating rate and growing importance of mass spectrometry applications to human blood proteins and peptides. Mass spectrometry can rapidly detect and identify the ionizable peptides from the proteins in a simple mixture and reveal many of their post-translational modifications. However, blood is a complex mixture that may contain many proteins first expressed in cells and tissues. The complete analysis of blood proteins is a daunting task that will rely on a wide range of disciplines from physics, chemistry, biochemistry, genetics, electromagnetic instrumentation, mathematics and computation. Therefore the comprehensive discovery and analysis of blood proteins will rank among the great technical challenges and require the cumulative sum of many of mankind's scientific achievements together. A variety of methods have been used to fractionate, analyze and identify proteins from blood, each yielding a small piece of the whole and throwing the great size of the task into sharp relief. The approaches attempted to date clearly indicate that enumerating the proteins and peptides of blood can be accomplished. There is no doubt that the mass spectrometry of blood will be crucial to the discovery and analysis of proteins, enzyme activities, and post-translational processes that underlay the mechanisms of disease. At present both discovery and quantification of proteins from blood are commonly reaching sensitivities of ∼1 ng/mL.

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“…There were a few publications using tandem high-performance chromatography mass spectrometry (LC-MS/MS) methods to quantify mAb (10)(11)(12)(13)(14)(15). The method approach involves enzyme digestion to reduce mAb into small peptides to be within the mass range of the mass spectrometry.…”

Section: Discussionmentioning

confidence: 99%

Bioanalytical Approaches to Quantify “Total” and “Free” Therapeutic Antibodies and Their Targets: Technical Challenges and PK/PD Applications Over the Course of Drug Development

Lee

Kelley

King

et al. 2011

AAPS J

244

151

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Abstract. The predominant driver of bioanalysis in supporting drug development is the intended use of the data. Ligand-binding assays (LBA) are widely used for the analysis of protein biotherapeutics and target ligands (L) to support pharmacokinetics/pharmacodynamics (PK/PD) and safety assessments. For monoclonal antibody drugs (mAb), in particular, which non-covalently bind to L, multiple forms of mAb and L can exist in vivo, including free mAb, free L, and mono-and/or bivalent complexes of mAb and L. Given the complexity of the dynamic binding equilibrium occurring in the body after dosing and multiple sources of perturbation of the equilibrium during bioanalysis, it is clear that ex vivo quantification of the forms of interest (free, bound, or total mAb and L) may differ from the actual ones in vivo. LBA reagents and assay formats can be designed in principle to measure the total or free forms of mAb and L. However, confirmation of the forms being measured under the specified conditions can be technically challenging. The assay forms and issues must be clearly communicated and understood appropriately by all stakeholders as the program proceeds through the development process. This paper focuses on monoclonal antibody biotherapeutics and their circulatory L that are either secreted as soluble forms or shed from membrane receptors. It presents an investigation into the theoretical and practical considerations for total/free analyte assessment to increase awareness in the scientific community and offer bioanalytical approaches to provide appropriate PK/PD information required at specific phases of drug development.

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Multiplexed Immunoassay: Quantitation and Profiling of Serum Biomarkers Using Magnetic Nanoprobes and MALDI-TOF MS

Cited by 77 publications

References 45 publications

Reference Measurement Procedure Development for C-Reactive Protein in Human Serum

Reference Measurement Procedure Development for C-Reactive Protein in Human Serum

Mass spectrometry of peptides and proteins from human blood

Bioanalytical Approaches to Quantify “Total” and “Free” Therapeutic Antibodies and Their Targets: Technical Challenges and PK/PD Applications Over the Course of Drug Development

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