Detection and Quantification of β-2-Microglobulin Using Mass Spectrometric Immunoassay

Tubbs, Kemmons A.; Nedelkov, Dobrin; Nelson, Randall W.

doi:10.1006/abio.2000.4921

Cited by 70 publications

(60 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Since its inception in 1995 by R.W. Nelson et al, 21 it has been applied to study a variety of proteins and peptides, [22][23][24][25][26][27][28][29][30][31][32][33][34][35][36] and has served as the foundation for the nascent field of Population Proteomics. [37][38][39][40][41][42] Roughly, MSIA can be envisioned as ultra high resolution, semiquantitative Western blotting in which protein variants are thoroughly resolved and their molecular masses determined to within less than 2 Da.…”

Section: Methodsmentioning

confidence: 99%

Glycosylation status of vitamin D binding protein in cancer patients

2009

Self Cite

View full text Add to dashboard Cite

On the basis of the results of activity studies, previous reports have suggested that vitamin D binding protein (DBP) is significantly or even completely deglycosylated in cancer patients, eliminating the molecular precursor of the immunologically important Gc macrophage activating factor (GcMAF), a glycosidase-derived product of DBP. The purpose of this investigation was to directly determine the relative degree of O-linked trisaccharide glycosylation of serumderived DBP in human breast, colorectal, pancreatic, and prostate cancer patients. Results obtained by electrospray ionization-based mass spectrometric immunoassay showed that there was no significant depletion of DBP trisaccharide glycosylation in the 56 cancer patients examined relative to healthy controls. These results suggest that alternative hypotheses regarding the molecular and/or structural origins of GcMAF must be considered to explain the relative inability of cancer patient serum to activate macrophages.

show abstract

Section: Methodsmentioning

confidence: 99%

Glycosylation status of vitamin D binding protein in cancer patients

2009

Self Cite

View full text Add to dashboard Cite

show abstract

“…Affinity pipette tips were prepared as previously described (12). In brief, frits were produced in bulk by loading soda lime glass beads into stainless steel annealing molds and baking them to form a solid, yet porous frit.…”

Section: Affinity Tip Manufacturementioning

confidence: 99%

“…As little as 50 l of whole blood was required for the sequential analysis of all three apolipoproteins from a single sample, with each assay taking as little as 15 min to perform (from sample collection to acquisition of spectra). The speed of the method relies on the development of affinity pipette tips (12), which utilize antibodies immobilized to a porous frit, for the isolation of the apolipoproteins from plasma. The availability of affinity pipette tips with large surface areas and high flow characteristics contribute to short incubation times without sacrificing sensitivities (detection on the order of femtomoles to picomoles).…”

Section: Protocol Development and Speed Of Analysismentioning

confidence: 99%

“…In this study, the use of mass spectrometric immunoassay (MSIA) (10)(11)(12) for structural characterization of apolipoproteins, apoA-I, apoA-II, and apoE was investigated. Important to the success of the MSIA approach were: 1 ) the development of devices and methods for the selective retrieval and preparation of the apolipoproteins for MALDI-TOF MS analysis; 2 ) the generation of high-quality mass spectrometric data capable of discerning variants of the apolipoproteins; and 3 ) the use of the approach in screening a small number of individuals in order to determine apolipoprotein profiles and, critically, in the phenotyping of apoE.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Novel mass spectrometric immunoassays for the rapid structural characterization of plasma apolipoproteins

Niederkofler

Tubbs

Kiernan

et al. 2003

Journal of Lipid Research

Self Cite

View full text Add to dashboard Cite

Novel mass spectrometric immunoassays (MSIAs) for the isolation and structural characterization of plasma apolipoprotein A-I (apoA-I), apoA-II, and apoE have been developed. The assays combine selective isolation of apolipoprotein species via affinity capture with mass-specific detection using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. In application, plasma (from 50 l of whole blood drawn from individuals, using finger lancet) was addressed with affinity pipette tips derivatized with antibodies toward the specific apolipoprotein. The time required for each assay was ‫ف‬ 15 min, less if assays on multiple individuals were performed in parallel. In a brief study of five individuals, several recently reported apoA-II variants were identified and observed consistently in all individuals. Additionally, the apoE phenotype of E3/ E3 was observed in three of the individuals, and E2/E3 and E3/E4 observed in the remaining two individuals, the latter of whom suffers from Alzheimer's disease. Overall, the MSIA approach offers a rapid, sensitive, and highly accurate means of profiling apolipoproteins from small volumes of plasma. The apolipoproteins represent a significant class of proteins whose functions range from enzyme activation (e.g., LCAT and lipoprotein lipase) to lipid/cholesterol transport and clearance. Although well studied over the past two decades, it has only recently become clear that mutations present within the apolipoproteins may be responsible for, or assist in, the progression of disease. Table 1 gives a brief overview of some diseases correlated with specific mutations present within apolipoproteins. Universally, these mutations are commonly studied on the DNA level using a variety of genotyping assays. For instance, apolipoprotein E (apoE) genotyping usually involves PCR amplification of short subsequences of exons 2 and 3 (containing both polymorphic sites) and analysis of the PCR products by: 1 ) dot blot hybridization with allele-specific oligonucleotide probes (1); 2 ) oligonucleotide ligation assay (2); 3 ) single-stranded conformational polymorphisms (3); or, 4 ) restriction fragment-length polymorphism analysis using restriction enzymes recognizing the sites of polymorphism (4). More advanced approaches involve competitive amplified single mutation detection by selective probe hybridization immunoassay (5) or primer-extension mass spectrometry assays directed at the polymorphic sites within the gene (6). This latter assay brings to the analysis the direct detection of indicators (extended primers containing the base complement of the polymorphism) at exact mass values, thus enabling an assay that is free of artifacts encountered in other assays using, e.g., electrophoretic gel shifts or differential hybridization.As with the nucleic acid mass spectrometry assays, direct mass spectrometric analysis of proteins (or proteolytic fragments) brings an unprecedented degree of analytical accuracy to the analysis of point mutations. However, whereas the polym...

show abstract

“…Work over the past 2 decades has led to the development of high-throughput mass spectrometric immunoassay (MSIA) 2 technologies and workflows that are well suited to the standardized characterization of protein diversity in human populations (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14). Data from such "population proteomics" studies include changes in concentration, amino acid sequence, and relative qualitative/quantitative changes in posttranslational modifications (15,16 ).…”

mentioning

confidence: 99%

Full-Length Characterization of Proteins in Human Populations

et al. 2010

View full text Add to dashboard Cite

BACKGROUND: Diversity in human proteins often gives rise to pluralities of structurally similar but functionally distinct proteins. Such microheterogeneity generally escapes proteomics discovery technologies, as well as conventional immunometric assays. As an intermediate between these 2 technological approaches, targeted, full-length characterization of proteins using mass spectrometry is a suitable means of defining microheterogeneity evident in human populations. CONTENT:We describe and explore the implications of microheterogeneity using the exemplar of human vitamin D binding protein (Gc-Globulin) as observed in cohorts of 400 individuals. Our investigations yielded: (a) population frequency data comparable to genotyping; (b) population frequency data for protein variants, with and without genotype linkage; (c) reference values for the different protein variants per cohort and genotype; and (d) associations between variant, frequency, relative abundance, and diseases.

show abstract

Detection and Quantification of β-2-Microglobulin Using Mass Spectrometric Immunoassay

Cited by 70 publications

References 22 publications

Glycosylation status of vitamin D binding protein in cancer patients

Glycosylation status of vitamin D binding protein in cancer patients

Novel mass spectrometric immunoassays for the rapid structural characterization of plasma apolipoproteins

Full-Length Characterization of Proteins in Human Populations

Contact Info

Product

Resources

About