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Research Articles

Facilitating the Validation of Novel Protein Biomarkers for Dementia (del Campo, Front. Neurol. 2015)


Different neurodegenerative disorders, such as Alzheimer’s disease (AD) and frontotemporal dementia (FTD), lead to dementia syndromes. Dementia will pose a hug impact on society and thus it is essential to develop novel tools that are able to detect th earliest, most sensitive, discriminative, and dynamic biomarkers for each of the disorders To date, the most common assays used in large-scale protein biomarker analysis ar enzyme-linked immunosorbent assays (ELISA), such as the sandwich immunoassays which are sensitive, practical, and easily implemented

The Interface Between Biomarker Discovery and Clinical Validation (Paulovich, Proteomics Clin Appl. 2008)


The application of “omics” technologies to biological samples generates hundreds to thousands of biomarker candidates; however, a discouragingly small number make it through the pipeline to clinical use. This is in large part due to the incredible mismatch between the large numbers of biomarker candidates and the paucity of reliable assays and methods for validation studies. We desperately need a pipeline that relieves this bottleneck between biomarker discovery and validation. This paper reviews the requirements for technologies to adequately credential biomarker candidates for costly clinical validation and proposes methods and systems to verify biomarker candidates. Models involving pooling of clinical samples, where appropriate, are discussed. We conclude that current proteomic technologies are on the cusp of significantly affecting translation of molecular diagnostics into the clinic.

Retraction: Wolfram Syndrome 1 and Adenylyl Cyclase 8 (Forseca, Nature Cell Biology, 2014)


Our Letter reported that WFS1 modulates insulin biosynthesis and secretion due to an interaction between WFS1 and AC8. It has recently become apparent that the anti-AC8 antibody, as well as some other key antibodies employed in the Letter are non-specific. In addition, we noted increased levels of WFS1 protein and the appearance of what may be post-translational modifications after incubation with high glucose (Fig. 5a,b and Supplementary Figs S4,S5).

Reproducibility Crisis: Blame It on the Antibodies (Baker, Nature, 2015)


In 2006, things were looking pretty good for David Rimm, a pathologist at Yale University in New Haven, Connecticut. He had developed a test to guide effective treatment of the skin cancer melanoma, and it promised to save lives. It relied on antibodies — large, Y-shaped proteins that bind to specified biomolecules and can be used to flag their presence in a sample. Rimm had found a combination of antibodies that, when used to 'stain' tumour biopsies, produced a pattern that indicated whether the patient would need to take certain harsh drugs to prevent a relapse after surgery. He had secured more than US$2 million in funding to move the test towards the clinic.

Practical Applications of Integrated Microfluidics for Peptide Quantification (Chambers, Bioanalysis. 2015)


Background: Increased pressure to obtain more, higher sensitivity data from less sample is especially critical for large peptides, whose already optimized LC–MS methods are heavily challenged by traditional ligand-binding assays.

Results: Critical bioanalytical assays were adapted to integrated microscale LC to reduce sample volumes while increasing sensitivity. Assays for teriparatide, glucagon and human insulin and five analogs were transferred from 2.1 mm analytical scale LC to a 150 µm scale system. This resulted in a 15–30 fold overall improvement in sensitivity derived from increased signal to noise, three to six fold reduction in injection volumes, and a two to five fold reduction in sample consumption.

Conclusion: Integrated microscale LC reduces sample consumption while enabling single picomolar quantification for therapeutic and endogenous peptides.

Improving the Measurement of Serum Thyroglobulin with Mass Spectrometry (Hoofnagle, JCEM, 2013)


Serum thyroglobulin (Tg) measurements are central to the management of patients treated for differentiated thyroid carcinoma. For decades, Tg measurements have relied on methods that are subject to interference by commonly found substances in human serum and plasma, such as Tg autoantibodies. As a result, many patients need additional imaging studies to rule out cancer persistence or recurrence that could be avoided with more sensitive and specific testing methods.

Ion Mobility Derived CCS to Support Metabolomics Applications (Paglia, Analytical Chem, 2015)


Metabolomics is a rapidly evolving analytical approach in life and health sciences. The structural elucidation of the metabolites of interest remains a major analytical challenge in the metabolomics workflow. Here, we investigate the use of ion mobility as a tool to aid metabolite identification. Ion mobility allows for the measurement of the rotationally averaged collision cross-section (CCS), which gives information about the ionic shape of a molecule in the gas phase.

The Bottleneck in the Cancer Biomarker Pipeline and Protein Quantification through Mass Spectrometry–Based Approaches: Current Strategies for Candidate Verification (Makawita, Clinical Chemistry 2009 )


Background: Although robust discovery-phase platforms have resulted in the generation of large numbers of candidate cancer biomarkers, a comparable system for subsequent quantitative assessment and verification of all candidates is lacking. Established immunoassays and available antibodies permit analysis of small subsets of candidates; however, the lack of commercially available reagents, coupled with high costs and lengthy production and purification times, have rendered the large majority of candidates untestable.

Application Notes / Posters

High-Throughput LC-MS MRM Disease Protein Marker Verification Using the ionKey/MS System (Hughes, Waters 2016 App note)


Biomarker discovery and validation are the first two steps in understanding disease and drug development. Validation is technology-challenged since it requires analyzing a large number of samples with high-throughput, but nevertheless requires high sensitivity, high resolution, large dynamic range and excellent selectivity.

A Comparison of Six Commercial Cortisol IA With LCMS (ARUP Poster, AACC, 2013)


Background: Measurement of serum cortisol is of particular importance in patients who are critically ill. However, commercial cortisol immunoassays exhibit differences in specificity, often attributed to potential cross reactivity with steroids other than cortisol. It is likely that these steroids may be elevated in critically ill patients, thus increasing the described inter‐assay variability between standard immunoassays.

Increasing Sensitivity and Minimizing Sample Volume for the Quantification of Therapeutic and Endogenous Cyclic Peptides in Plasma Using ionKey/MS (Lamb, Waters 2016 App note)


The use of peptides and proteins as therapeutic agents has increased significantly in recent years. Thus, the demand for their analysis is toxicokinetic and pharmacokinetic studies is increasing as well.

Ultrasensitive Quantification Assay for Oxytocin in Human Plasma Using the ionKey/MS System (Doneanu, Waters 2016 App note)


Quantitative LC-MRM (multiple reaction monitoring) methods for small molecule drugs are used to provide bioanalytical support in various stages of drug discovery and development. These methods can routinely achieve lower limits of quantification (LLOQ) in the range of 50 to 100pg/mL, in various biological matrices, using analytical-scale chromatography (e.g., 2.1 mm I.D. UPLC® Columns). In the case of peptide therapeutics, these assays are more challenging because lower LLOQs are often required.

Development of a Quantitative SPE/LC/MS/MS Assay for Multiple Amyloid Beta Peptide in Cerebrospinal Fluid (Chambers, ASMS 2010 Poster)


The deposition/formation of insoluble aggregates of amyloid β peptides (Aβ) in the brain is considered to be a critical event in Alzheimer's Disease (AD). Therapeutic strategies have focused on small molecule inhibitors or enhancing its clearance. Therefore, the need for highly sensitive and robust methods for quantitation of amyloid peptides in cerebrospinal fluid and their relationship to AD is of critical interest to many researchers. However, the analysis of these Aβ peptides is extremely challenging due not only to the relatively low abundance in biological fluids but also because they have a propensity for aggregation, they are potentially bound by other proteins and have the tendency to form oligomers.

Case Studies / Webinars

LC-MS for Food Science


Dr. Jessica Prenni and her team at Colorado State University's (CSU) Proteomics and Metabolomics Facility discuss how the next discovery in food science may come from the field of metabolomics and the significant role the ionKey/MS system is playing.

LC-MS for Tumor Associated Antigens (TAA)


Duke University’s Proteomics and Metabolomics Core Facility details how Waters’ ionKey/MS technology quantifies tumor associated antigens, and the impact this could have on healthcare.

White Papers

Taking Advantage of Significant Reductions in Ion Suppression Using ionKey/MS Compared to Standard-Flow LC-MS (Johnson, Waters White Paper)


This white paper compares ion suppression levels of the Waters ionKey/MS™ System to those of a standardflow LC/MS system. In both peptide and small molecule applications, it is shown that the ionKey/MS System delivers a significant reduction in ion suppression characteristics due to the lower flow rates utilized during its analysis.

Enhancing Mass Spectrometry Sensitivity By Reducing Chromatographic Flow Rates with ionKey/MS (Murphy, Waters White Paper)


This white paper will review the sensitivity benefits that can be expected when operating at a microscale flow rate with the Waters ionKey/MS™ System in comparison to a 2.1-mm column format, as well as explain why this signal enhancement is possible.

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