• Home
  • Norman Leigh Anderson
Norman Leigh Anderson

Norman Leigh Anderson
SISCAPA Assay Technologies

PhD

About

311
Publications
16,660
Reads
How we measure 'reads'
A 'read' is counted each time someone views a publication summary (such as the title, abstract, and list of authors), clicks on a figure, or views or downloads the full-text. Learn more
28,819
Citations

Publications

Publications (311)
Preprint
INTRODUCTION The pandemic readiness toolbox needs to be extended, providing diagnostic tools that target different biomolecules, using orthogonal experimental setups and fit-for-purpose specification of detection. Here we build on a previous Cov-MS effort that used liquid chromatography-mass spectrometry (LC-MS) and describe a method that allows ac...
Preprint
Full-text available
Reliable, robust, large-scale molecular testing for SARS-CoV-2 is essential for monitoring the ongoing Covid-19 pandemic. We have developed a scalable analytical approach to detect viral proteins based on peptide immunoaffinity enrichment combined with liquid chromatography - mass spectrometry (LC-MS). This is a multiplexed strategy, based on targe...
Preprint
Full-text available
High-frequency longitudinal tracking of inflammation using dried blood microsamples provides a new window for personalized monitoring of infections, chronic inflammatory disease, and clinical trials of anti-inflammatory drugs. Using 1,662 dried blood spot samples collected by 16 subjects over periods of weeks to years, we studied the behavior of 12...
Conference Paper
Full-text available
Background Current serology-based, treponemal-specific diagnostic tests detect antibodies reactive against T. pallidum molecules and cannot differentiate between past and current syphilis infections. Further, existing diagnostic tests for syphilis have non-optimal sensitivity and specificity and require expertise for test administration and interpr...
Preprint
Full-text available
A detailed understanding of changes in blood protein biomarkers occuring in individuals over time would enable truly personalized approaches to health and disease monitoring. Such measurements could reveal smaller, earlier departures from normal baseline levels of biomarkers thus allowing better disease detection and treatment monitoring. Current p...
Article
Aim: Hybrid LC-MS/MS assays are increasingly used to quantitate proteins in biological matrices. These assays involve analyte enrichment at the protein level. Although suitability has been demonstrated, they are limited by the lack of appropriate affinity reagents and may suffer from interferences caused by binding proteins or antibodies. Results...
Article
Background: Lipoprotein-associated phospholipase A2 (Lp-PLA2), an enzyme associated with inflammation, is used as a biomarker for cardiovascular disease risk. Both the concentration and activity of Lp-PLA2 have been shown to be clinically relevant. However, there is a discordance between the serum concentration of Lp-PLA2 measured by the standard...
Article
Background: The use of DBS for quantitative protein biomarker measurement has been hindered by issues associated with blood hematocrit variations and lack of detection sensitivity, particularly when multiple biomarkers are measured. Materials & methods: An automated, multiplexed SISCAPA analysis was used to normalize blood volume variations in D...
Article
Introduction: Aided by the advent of advanced mass spectrometry (MS)-based technologies and methodologies, quantitative proteomics has emerged as a viable technique to capture meaningful data for candidate biomarker evaluation. Areas covered: To aid clinical translation, these methods generally utilize a bottom-up strategy with isotopically labe...
Article
Full-text available
Almost all clinically used protein biomarkers are currently measured by individual immunoassays on separate sample aliquots, an approach that entails penalties in terms of cost and sample consumption for each additional analyte measured. Thus, if an additional biomarker contributes less than a startling increase in predictive power, or even if it p...
Article
Full-text available
Context: Prostate-specific antigen (PSA) is a 34-kDa glycoprotein with chymotrypsin-like enzyme activity that circulates both in free forms and complexed to various enzyme inhibitors including antichymotrypsin and α2-macroglobulin. Prostate-specific antigen bound to α2-macroglobulin is not detected by commercial PSA immunoassays. Objective: To d...
Article
The eighth Workshop on Recent Innovations in Bioanalysis (WRIB) drew close to 500 professionals representing large pharmas, biotechs, CROs and multiple regulatory agencies from around the world, working on both small and large molecule bioanalysis. This year, Bioanalysis was proud to support the first WRIB Poster Award. Posters were judged by an ex...
Article
Full-text available
Objectives: Harmonization of prostate-specific antigen (PSA) immunoassays is important for good patient care. The specificity of the antibodies used to detect circulating PSA could cause differences in the PSA measurements. Methods: We used mass spectrometry (MS) to quantitate the concentration of five peptides cleaved from trypsin digestion of...
Article
This review describes one thread in a fabric of developments leading to the present state of proteomics, stretching over 60years and ending with a prediction for 2024. While composed largely of personal reminiscences, the story offers some instructive successes and failures, and appears to be nearing the long-sought goal of deep insights into real...
Article
Full-text available
Adoption of targeted mass spectrometry (MS) approaches such as multiple reaction monitoring (MRM) to study biological and biomedical questions is well underway in the proteomics community. Successful application depends on the ability to generate reliable assays that uniquely and confidently identify target peptides in a sample. Unfortunately, ther...
Article
Full-text available
Protein biomarkers are needed to deepen our understanding of cancer biology and to improve our ability to diagnose, monitor and treat cancers. Important analytical and clinical hurdles must be overcome to allow the most promising protein biomarker candidates to advance into clinical validation studies. Although contemporary proteomics technologies...
Article
Full-text available
Background: Biomarker validation remains one of the most challenging constraints to the development of new diagnostic assays. To facilitate biomarker validation, we previously developed a chromatography-free stable isotope standards and capture by antipeptide antibodies (SISCAPA)-MALDI assay allowing rapid, high-throughput quantification of protei...
Article
> In theory, there is no difference between theory and practice. In practice, there is. > > —variously attributed to Andrew Tannenbaum, Jan L.A. van de Snepscheut, or Yogi Berra The theory behind an exploding kaleidoscope of protein biomarker research is that reproducible molecular differences that correlate with important medical attributes can b...
Article
We investigated the utility of an SPE-MS/MS platform in combination with a modified SISCAPA workflow for chromatography-free MRM analysis of proteotypic peptides in digested human plasma. This combination of SISCAPA and SPE-MS/MS technology allows sensitive, MRM-based quantification of peptides from plasma digests with a sample cycle time of ~7 sec...
Article
We have investigated the precision of peptide quantitation by MALDI-TOF mass spectrometry (MS) using six pairs of proteotypic peptides (light) and same-sequence stable isotope labeled synthetic internal standards (heavy). These were combined in two types of dilution curves spanning 100-fold and 2000-fold ratios. Coefficients of variation (CV; stand...
Article
Full-text available
The inability to quantify large numbers of proteins in tissues and biofluids with high precision, sensitivity, and throughput is a major bottleneck in biomarker studies. We previously demonstrated that coupling immunoaffinity enrichment using anti-peptide antibodies (SISCAPA) to multiple reaction monitoring mass spectrometry (MRM-MS) produces Immun...
Article
Full-text available
There is a great need for quantitative assays in measuring proteins. Traditional sandwich immunoassays, largely considered the gold standard in quantitation, are associated with a high cost, long lead time, and are fraught with drawbacks (e.g. heterophilic antibodies, autoantibody interference, 'hook-effect').(1) An alternative technique is affinit...
Article
Many investigators have used proteomic methodologies to investigate specific questions in basic biology and medical science. Indeed, many millions of dollars have been spent around the world on the generation of extensive lists of proteins that reside in a particular organelle, cell type, tissue, or body fluid. These lists represent the proteomes o...
Article
Full-text available
Stable isotope standards and capture by antipeptide antibodies (SISCAPA) couples affinity enrichment of peptides with stable isotope dilution and detection by multiple reaction monitoring mass spectrometry to provide quantitative measurement of peptides as surrogates for their respective proteins. In this report, we describe a feasibility study to...
Article
Clinical proteomics presents great promise in biology and medicine because of its potential for improving our understanding of diseases at the molecular level and for detecting disease-related biomarkers for diagnosis, prognosis, and prediction of therapeutic responses. To realize its full potential to improve clinical outcome for patients, proteom...
Article
Full-text available
Featured Article: Anderson NL, Polanski M, Pieper R, Gatlin T, Tirumalai RS, Conrads TP, et al. The human plasma proteome: a non-redundant list developed by combination of four separate sources. Mol Cell Proteomics 2004;3:311–26.2 The problem of enumerating protein components in plasma has challenged the best analytical technologies for more than...
Article
A scalable method for screening and selection of peptide-specific monoclonal antibodies (mAbs) is described. To identify high affinity anti-peptide mAbs in hybridoma supernatants, antibodies were captured by magnetic affinity beads followed by binding of specific peptides from solution. After timed washing steps, the remaining bound peptides were e...
Article
Although only approximately 200 protein analytes, equivalent to 1% of the human proteome, are currently measured in plasma or serum for clinical purposes, there is an enormous unmet need for new or improved clinical diagnostic tests in many disease areas. Substantial biomarker discovery efforts over the last 5 years have generated hundreds—in some...
Article
Full-text available
This review reports on the current and emerging technologies for the use of mass-spectrometry-based proteomics in clinical applications.
Article
Cancer has profound effects on gene expression, including a cell's glycosylation machinery. Thus, tumors produce glycoproteins that carry oligosaccharides with structures that are markedly different from the same protein produced by a normal cell. A single protein can have many glycosylation sites that greatly amplify the signals they generate comp...
Article
Full-text available
As a part of ongoing efforts of the NCI-FDA Interagency Oncology Task Force subcommittee on molecular diagnostics, members of the Clinical Proteomic Technology Assessment for Cancer program of the National Cancer Institute have submitted 2 protein-based multiplex assay descriptions to the Office of In Vitro Diagnostic Device Evaluation and Safety,...
Article
Full-text available
Clinical proteomics has the potential to enable the early detection of cancer through the development of multiplex assays that can inform clinical decisions. However, there has been some uncertainty among translational researchers and developers as to the specific analytical measurement criteria needed to validate protein-based multiplex assays. To...
Article
Full-text available
An analysis of all US Food and Drug Administration (FDA) approvals for protein-based assays through 2008 reveals 109 unique protein targets in plasma or serum, as well as 62 additional tests for peptides, protein posttranslational modifications, protein complexes, autoantibodies against endogenous proteins, and blood cell proteins. A further 96 uni...
Article
Full-text available
“ The determination of the structure of insulin clearly opens up the way to similar studies on other proteins … One may also hope that studies on proteins may reveal changes that take place in disease and that our efforts may be of more practical use to humanity. ” — Frederick S. Sanger, Nobel Lecture December 11, 1958 (1) This special issue of C...
Article
Full-text available
There is an urgent need for quantitative assays in verifying and validating the large numbers of protein biomarker candidates produced in modern "-omics" experiments. Stable isotope standards with capture by anti-peptide antibodies (SISCAPA) has shown tremendous potential to meet this need by combining peptide immunoaffinity enrichment with quantit...
Article
Full-text available
Verification of candidate biomarkers relies upon specific, quantitative assays optimized for selective detection of target proteins, and is increasingly viewed as a critical step in the discovery pipeline that bridges unbiased biomarker discovery to preclinical validation. Although individual laboratories have demonstrated that multiple reaction mo...
Article
Full-text available
Mass spectrometry-based multiple reaction monitoring (MRM) quantitation of proteins can dramatically impact the discovery and quantitation of biomarkers via rapid, targeted, multiplexed protein expression profiling of clinical samples. A mixture of 45 peptide standards, easily adaptable to common plasma proteomics work flows, was created to permit...
Article
Full-text available
A SISCAPA (stable isotope standards and capture by anti-peptide antibodies) method for specific antibody-based capture of individual tryptic peptides from a digest of whole human plasma was developed using a simplified magnetic bead protocol and a novel rotary magnetic bead trap device. Following off-line equilibrium binding of peptides by antibodi...
Article
Full-text available
The lack of sensitive, specific, multiplexable assays for most human proteins is the major technical barrier impeding development of candidate biomarkers into clinically useful tests. Recent progress in mass spectrometry-based assays for proteotypic peptides, particularly those with specific affinity peptide enrichment, offers a systematic and econ...
Article
Full-text available
The enormous potential of biomarkers to revolutionize clinical practice and improve patient care has been well documented (1)(2). Molecular-based diagnostic and prognostic tests, particularly those aimed at protein analytes, could be used to detect disease earlier, enabling treatment to start sooner and possibly cure rather than to merely delay fur...
Article
A refined surface plasmon resonance method was developed to measure the kinetics of peptide binding to rabbit monoclonal antibodies (RabMAbs). Optimized amounts of RabMAbs were captured onto sensor chips from hybridoma supernatants followed by binding of free peptides from solution. This allowed kinetic measurement of monovalent interactions of pep...
Article
Full-text available
Plasma contains thousands of proteins, but a small number of these proteins comprise the majority of protein molecules and mass. We surveyed proteomic studies to identify candidates for high-abundance polypeptide chains. We searched the literature for information on the plasma concentrations of the most abundant components in healthy adults and for...
Article
Full-text available
the MIAPE Gel Electrophoresis (MIAPE-GE) guidelines specify the minimum information that should be provided when reporting the use of n-dimensional gel electrophoresis in a proteomics experiment. Developed through a joint effort between the gel-based analysis working group of the Human Proteome Organisation's Proteomics Standards Initiative (HUPO-P...
Article
Full-text available
CLUB ("Candidate List of yoUr Biomarkers") is a freely available, web-based resource designed to support Cancer biomarker research. It is targeted to provide a comprehensive list of candidate biomarkers for various cancers that have been reported by the research community. CLUB provides tools for comparison of marker candidates from different exper...
Article
Reliable study results are necessary for the assessment of discoveries, including those from proteomics. Reliable study results are also crucial to increase the likelihood of making a successful choice of biomarker candidates for verification and subsequent validation studies, a current bottleneck for the transition to in vitro diagnostic (IVD). In...
Article
A major bottleneck for validation of new clinical diagnostics is the development of highly sensitive and specific assays for quantifying proteins. We previously described a method, stable isotope standards with capture by antipeptide antibodies, wherein a specific tryptic peptide is selected as a stoichiometric representative of the protein from wh...
Article
Full-text available
Multiple approaches for simplifying the serum proteome have been described. These techniques are generally developed across different laboratories, samples, mass spectrometry platforms, and analysis tools. Hence, comparing the available schemes is impossible from the existing literature because of confounding variables. We describe a head-to-head c...
Article
Full-text available
We have compiled from literature and other sources a list of 1261 proteins believed to be differentially expressed in human cancer. These proteins, only some of which have been detected in plasma to date, represent a population of candidate plasma biomarkers that could be useful in early cancer detection and monitoring given sufficiently sensitive...
Article
In any attempt to construct a catalog of the proteins of a given species, the genetic heterogeneity of natural plant and animal populations makes it necessary to consider variants of each protein. Thus in compiling a Human Protein Index using high resolution two-dimensional electrophoresis as a separating technique, protein variants differing from...
Article
Full-text available
On the basis of discussions with representatives from all sectors of the cancer research community, the National Cancer Institute (NCI) recognizes the immense opportunities to apply proteomics technologies to further cancer research. Validated and well characterized affinity capture reagents (e.g. antibodies, aptamers, and affibodies) will play a k...
Article
Full-text available
Quantitative LC-MS/MS assays were designed for tryptic peptides representing 53 high and medium abundance proteins in human plasma using a multiplexed multiple reaction monitoring (MRM) approach. Of these, 47 produced acceptable quantitative data, demonstrating within-run coefficients of variation (CVs) (n = 10) of 2-22% (78% of assays had CV <10%)...
Article
Full-text available
Numerous recent reports (e.g. Refs. 1-3) suggest that pro- teome studies are in the process of finding a range of novel disease biomarkers in plasma and serum, giving rise to the hope that the declining trend in new protein diagnostics over the last decade (4) will be reversed and the obvious benefits of early (5) and correct diagnosis will be exte...
Article
For the translation of marker candidates and technologies from proteomics into in vitro diagnostic (IVD) tests, IVD requirements and needs have to be considered explicitly. Subject to stringent cost/benefit analyses, IVDs have to provide (1) definite and reliable diagnosis tied to decisions on interventions, (2) adequate test accuracy (bias and imp...
Article
Biomarkers for cancer risk, early detection, prognosis, and therapeutic response promise to revolutionize cancer management. Protein biomarkers offer tremendous potential in this regard due to their great diversity and intimate involvement in physiology. An effective program to discover protein biomarkers using existing technology will require team...
Article
The key concept of proteomics (looking at many proteins at once) opens new avenues in the search for clinically useful biomarkers of disease, treatment response and ageing. As the number of proteins that can be detected in plasma or serum (the primary clinical diagnostic samples) increases towards 1000, a paradoxical decline has occurred in the num...
Article
Full-text available
We have merged four different views of the human plasma proteome, based on different methodologies, into a single nonredundant list of 1175 distinct gene products. The methodologies used were 1) literature search for proteins reported to occur in plasma or serum; 2) multidimensional chromatography of proteins followed by two-dimensional electrophor...
Article
The National Heart, Lung, and Blood Institute (NHLBI) Clinical Proteomics Working Group was charged with identifying opportunities and challenges in clinical proteomics and using these as a basis for recommendations aimed at directly improving patient care. The group included representatives of clinical and translational research, proteomic technol...
Article
To facilitate the construction, functional characterization, and use of immunoadsorbents, we have developed a flow cytometry method that allows rapid assessment of large numbers of particle-bound antibodies. Protein G derivitized POROS beads were used to bind affinity-purified antibodies specific for synthetic peptides designed from human plasma pr...
Article
Full-text available
A method (denoted SISCAPA) for quantitation of peptides in complex digests is described. In the method, anti-peptide antibodies immobilized on 100 nanoliter nanoaffinity columns are used to enrich specific peptides along with spiked stable-isotope-labeled internal standards of the same sequence. Upon elution from the anti-peptide antibody supports,...
Article
Full-text available
We propose a system for continuing surveillance of viral pathogens circulating in large human populations. We base this system on the physical isolation of viruses from large pooled samples of human serum and plasma (e.g., discarded specimens from diagnostic laboratories), followed by shotgun sequencing of the resulting genomes. The technology for...
Article
Plasma, the soluble component of the human blood, is believed to harbor thousands of distinct proteins, which originate from a variety of cells and tissues through either active secretion or leakage from blood cells or tissues. The dynamic range of plasma protein concentrations comprises at least nine orders of magnitude. Proteins involved in coagu...
Article
Plasma contains numerous and diverse proteins with existing and potential therapeutic value. Plasma has been used clinically as both a source of purified derivatives for treating diseases such as hemophilia, and as a diagnostic medium. Recent research directed towards mining plasma's true potential takes advantage of state-of-the-art proteomic anal...
Article
In order to discover novel protein markers indicative of disease processes or drug effects, the proteomics technology platform most commonly used consists of high resolution protein separation by two-dimensional electrophoresis (2-DE), mass spectrometric identification of proteins from stained gel spots and a bioinformatic data analysis process sup...