Enzyme Immunoassay and Enzyme-Linked Immunosorbent Assay

Gan, Stephanie D.; Patel, Kruti

doi:10.1038/jid.2013.287

Cited by 449 publications

(291 citation statements)

References 6 publications

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“…Many specific serum proteins can also be measured by immunoassays, which require a specific antibody against the analysed serum protein. Enzyme immunoassay and enzyme-linked immunosorbent assay (ELISA) are the most common analytical tools in biomedical research for the detection and quantification of specific proteins, antigens or antibodies (Gan and Patel 2013). ELISA is based on the concept of an antigen/protein binding to its specific antibody, which allows detection of very low concentrations of antigen/protein (Mariani et al 1998).…”

Section: Analysis Of Specific Serum Proteinsmentioning

confidence: 99%

Serum proteins and their diagnostic utility in veterinary medicine: a review

Tóthová¹,

Nagy²,

Kováč³

2016

Vet. Med.

134

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ABSTRACT:The evaluation of serum proteins and their electrophoretic pattern is a well-established laboratory method in the diagnosis of many diseases in humans, which has replaced the biochemical determination of the concentrations of albumin and the ratio of albumin to globulins. The measurement of serum proteins may be an important diagnostic tool for the detection, diagnosis, and monitoring of various diseases and pathological processes. The results of serum protein electrophoresis can be one of the most useful diagnostic aids in a wide spectrum of diseases, including infectious and inflammatory diseases, renal or gastrointestinal disorders, immunodeficiency states, as well as paraproteinaemias caused by lymphoid or plasma cell neoplasia. Although many studies have been carried out to determine the usefulness of the determination of serum proteins and their electrophoretic pattern in various disease conditions and disorders also in animals, serum protein evaluation is still a relatively little-used diagnostic tool in veterinary medicine. In this article, methods of serum protein determination, their diagnostic utility in animal care practice and their different patterns in dysproteinaemias and paraproteinaemias are reviewed.

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Section: Analysis Of Specific Serum Proteinsmentioning

confidence: 99%

Serum proteins and their diagnostic utility in veterinary medicine: a review

Tóthová¹,

Nagy²,

Kováč³

2016

Vet. Med.

134

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“…An antibody against the target protein is coated in the wells; the crude sample is added and incubated to allow the target protein to bind and a primary antibody that recognizes the target protein but at a different epitope is added. In a sandwich-ELISA, this primary antibody can either be labeled with an enzyme or a secondary labeled antibody is needed (210). In paper I, a colorimetric sandwich-ELISA was used.…”

Section: Elisamentioning

confidence: 99%

Understanding the dual nature of lysozyme: part villain – part hero : A Drosophila melanogaster model of lysozyme amyloidosis

Helmfors¹

2014

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Amyloid proteins are a distinct class of proteins that can misfold into β-sheet rich structures that later mature to form the characteristic species known as amyloid fibrils, and accumulate in tissues in the human body. The misfolding event is often caused by mutations (or outer factors such as changes in pH) that destabilize the native protein structure. The mature amyloid fibrils were initially believed to be associated with diseases connected to protein misfolding such as Alzheimer's disease (AD), Parkinson's disease, transthyretin amyloidosis and lysozyme amyloidosis. However, now it is known that many different factors are involved in these diseases such as failure in protein clearance, lysosomal dysfunction and formation of intermediate misfolded protein species, which possess cytotoxic properties, preceding the formation of mature fibrils.In this thesis the amyloidogenic protein lysozyme has been examined in vivo by using Drosophila melanogaster (fruit fly) as a model organism. The effects of over-expressing human lysozyme and amyloidogenic variants in Drosophila have been investigated both in the absence and presence of the serum amyloid P component (SAP), a protein known to interact with amyloid species. In addition, the role of lysozyme in AD has been investigated by co-expressing human lysozyme and amyloid β in Drosophila.The lysozyme protein is an enzyme naturally found in bodily fluids such as tears, breast milk and saliva. It is engaged in the body's defense and acts by hydrolyzing the cell wall of invading bacteria. Certain disease-associated point mutations in the gene encoding lysozyme destabilize the protein and cause it to misfold which results in systemic amyloidosis. To investigate the in vivo misfolding behavior of lysozyme we developed and established a Drosophila model of lysozyme amyloidosis. SAP is commonly found attached to amyloid deposits in the body; however, the role of SAP in amyloid diseases is unknown. To investigate the effect of SAP in lysozyme misfolding, these two proteins were co-expressed in Drosophila.The amyloid β peptide is involved in AD, building up the plaques found in AD patient brains. These plaques trigger neuroinflammation and since lysozyme is upregulated during various inflammation conditions, a possible role of lysozyme in AD was investigated by overexpressing lysozyme in a Drosophila model of AD. Interaction between lysozyme and the amyloid β protein was also studied by biophysical measurements.During my work with this thesis, the dual nature of lysozyme emerged; on the one hand a villain, twisted by mutations, causing the lysozyme amyloidosis disease. On the other hand a hero, delaying the toxicity and maybe the neurological damage caused by the amyloid β peptide.iii Populärvetenskaplig sammanfattningProteiner är viktiga byggstenar i naturen. De bygger upp hud, hår, muskler och organ, påskyndar kemiska reaktioner i kroppen och transporterar till exempel syre och andra molekyler i kroppen. Proteiner byggs upp av aminosyror, liksom pärlor på ett halsba...

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“…While the protein quantitation strategies are now wellestablished, a unique challenge for reliable and sensitive localization and quantitation of protein therapeutics based on or mimicking abundant endogenous proteins (such as Hp) arises from the fact that such measurements are carried out on the varying (and often unpredictable) background of endogenous molecules, the structure of which is identical to that of the exogenous (administered) proteins. This makes it highly problematic to use classic techniques of protein quantitation, such as enzyme-linked immunosorbent assay (ELISA) [16,17], which remains one of the most popular and sensitive techniques in the field [18,19]. Recently, mass spectrometry (MS) emerged as a powerful tool for protein quantitation in complex biological matrices, enabling highly sensitive and selective measurements both in the field of proteomics [20] and in a variety of pharmacokinetic studies [21][22][23].…”

Section: Introductionmentioning

confidence: 99%

Evaluation of Gallium as a Tracer of Exogenous Hemoglobin–Haptoglobin Complexes for Targeted Drug Delivery Applications

Kaltashov

2016

J. Am. Soc. Mass Spectrom.

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Abstract. Haptoglobin (Hp) is a plasma glycoprotein that generates significant interest in the drug delivery community because of its potential for delivery of antiretroviral medicines with high selectivity to macrophages and monocytes, the latent reservoirs of human immunodeficiency virus. As is the case with other therapies that exploit transport networks for targeted drug delivery, the success of the design and optimization of Hp-based therapies will critically depend on the ability to accurately localize and quantitate Hp-drug conjugates on the varying and unpredictable background of endogenous proteins having identical structure. In this work, we introduce a new strategy for detecting and quantitating exogenous Hp and Hp-based drugs with high sensitivity in complex biological samples using gallium as a tracer of this protein and inductively coupled plasma mass spectrometry (ICP MS) as a method of detection. Metal label is introduced by reconstituting hemoglobin (Hb) with gallium(III)-protoporphyrin IX followed by its complexation with Hp. Formation of the Hp/Hb assembly and its stability are evaluated with native electrospray ionization mass spectrometry. Both stable isotopes of Ga give rise to an abundant signal in ICP MS of a human plasma sample spiked with the metallabeled Hp/Hb complex. The metal label signal exceeds the spectral interferences' contributions by more than an order of magnitude even with the concentration of the exogenous protein below 10 nM, the level that is more than adequate for the planned pharmacokinetic studies of Hp-based therapeutics.

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Enzyme Immunoassay and Enzyme-Linked Immunosorbent Assay

Cited by 449 publications

References 6 publications

Serum proteins and their diagnostic utility in veterinary medicine: a review

Serum proteins and their diagnostic utility in veterinary medicine: a review

Understanding the dual nature of lysozyme: part villain – part hero : A Drosophila melanogaster model of lysozyme amyloidosis

Evaluation of Gallium as a Tracer of Exogenous Hemoglobin–Haptoglobin Complexes for Targeted Drug Delivery Applications

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