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Highly selective artificial gel antibodies for detection and quantification of biomarkers in clinical samples: I. Spectrophotometric approach to design the calibration curve for the quantification
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
2008 (English)In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 31, no 22, 3945-3953 p.Article in journal (Refereed) Published
Abstract [en]

High selectivity of a biomarker is a basic requirement when it is used for diagnosis, prognosis and treatment of a disease. The artificial gel antibodies, which we synthesise by a molecular imprinting method, have this property not only for proteins, but also for bioparticles, such as viruses and bacteria. However, diagnosis of a disease requires not only that the biomarker can be "fished out" from a body fluid with high selectivity, but also that its concentration in the sample can rapidly be determined and preferably by a simple technique. This paper deals primarily with the development of a spectrophotometric method, which is so simple and fast that it can be used with advantage in a Doctor's Office. The development of this method was not straight-forward. However, by modifications of the performance of these measurements we can now design standard curves in the form of a straight line, when we plot the true (not the recorded "apparent" absorption) against known protein concentrations. In an additional publication (see the following paper in this issue of JSS) we show an application of such a plot: determination of the concentration of albumin in serum and cerebrospinal fluid from patients with neurological disorders to investigate whether albumin is a biomarker for these diseases.

Place, publisher, year, edition, pages
2008. Vol. 31, no 22, 3945-3953 p.
Keyword [en]
Apparent absorption, artificial antibodies, CBB, molecular imprinting, true absorption
National Category
Pharmaceutical Sciences
Identifiers
URN: urn:nbn:se:uu:diva-104856DOI: 10.1002/jssc.200800385ISI: 000262167500017PubMedID: 19065619OAI: oai:DiVA.org:uu-104856DiVA: diva2:220189
Available from: 2009-05-29 Created: 2009-05-29 Last updated: 2017-12-13Bibliographically approved
In thesis
1. Application of Artificial Gel Antibodies for the Detection and Quantification of Proteins in Biological Fluids
Open this publication in new window or tab >>Application of Artificial Gel Antibodies for the Detection and Quantification of Proteins in Biological Fluids
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The molecular-imprinting method has previously been used for the synthesis of artificial gel antibodies, highly selective for various proteins. In present study, we have synthesized artificial gel antibodies against haemoglobin, albumin and different forms of growth hormone with the aim to develop a simple and rapid procedure to measure the concentration of these protein biomarkers in samples of clinical interest.  A spectrophotometric method was developed to design a standard curve in the form of a straight line, whereby the true absorption (not the recorded “apparent” absorption) was plotted against a known protein concentration. The procedure, applied to quantitative analysis of albumin in human plasma and cerebrospinal fluid (CSF) from patients with ALS, indicated that  the concentration of this protein was significantly enhanced in CSF from patients with amyotrophic lateral sclerosis (ALS), compared to control samples. A low level of albumin was observed in plasma from ALS patients compared to controls. Additionally, free zone electrophoresis was employed to detect human growth hormone (GH) activity in hormone preparations purified from human pituitaries. We have successfully synthesized antibodies capable of discriminating between dimeric and monomeric GH in samples of clinical origin. To quantify these proteins a calibration curve has been designed, i.e. a plot of the electrophoretic mobility of the complex GH/gel antibody against the protein concentration in the sample, for instance serum or CSF.

This method was also employed for qualitative and quantitative determinations of Somatropin, a non-glycosylated GH and glycosylated-GH in a body liquid.

Our results indicate that by this technique one can “fish out” with high accuracy various proteins from both body fluids containing a great number of other proteins. It might well apply also to biomarker proteins for other diseases.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2010. 64 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 741
National Category
Medicinal Chemistry Pharmaceutical Sciences
Research subject
Pharmaceutical Science
Identifiers
urn:nbn:se:uu:diva-122457 (URN)978-91-554-7802-5 (ISBN)
Public defence
2010-05-18, C4:305, Uppsala biomedicinska centrum BMC, Husarg. 3, Uppsala, 13:00 (English)
Opponent
Supervisors
Available from: 2010-04-27 Created: 2010-04-13 Last updated: 2010-05-17Bibliographically approved

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