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Applying hollow fibres for separating free and bound label in continuous-flow immunochemical detection
Division of Analytical Chemistry, Leiden/Amsterdam Center for Drug Research, Leiden University, Leiden, The Netherlands.
Division of Analytical Chemistry, Leiden/Amsterdam Center for Drug Research, Leiden University, Leiden, The Netherlands.
Division of Analytical Chemistry, Leiden/Amsterdam Center for Drug Research, Leiden University, Leiden, The Netherlands.
Division of Analytical Chemistry, Leiden/Amsterdam Center for Drug Research, Leiden University, Leiden, The Netherlands.
1996 (English)In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 755, no 2, 179-187 p.Article in journal (Refereed) Published
Abstract [en]

On-line liquid chromatography-immunochemical detection (LC-ICD) provides the possibility to individually monitor cross-reactive compounds overcoming the need of tedious fraction collection. ICD is performed as a post-column reaction detection system and is based on a two-step immunoreaction. In the first step unlabelled antibodies are added to the LC effluent and allowed to react with antigens (analytes) eluting from the LC column. The amount of analytes bound to the antibodies is measured by adding, in a second step, labelled antigen to the reaction mixture. For quantitation, free and bound label need to be separated prior to detection. The present paper describes a hollow fibre module (HFM), which can be used for this purpose. Separation of free and bound label occurs on discrimination by size. Using biotin as a model compound, a detection limit of 30 nmol/l can be reached employing anti-biotin antibodies and a low-molecular-mass fluorescence label in the LC-ICD system. Additional to low-molecular-mass labels, the HFM allows the use of small enzyme labels. In this context, horseradish peroxidase-labelled biotin was used as a label in combination with antibodies in the immunochemical detection of biotin. This allows future implementation of commercially available enzyme immunoassay kits in continuous-flow immunochemical detection.

Place, publisher, year, edition, pages
Amsterdam: Elsevier B.V , 1996. Vol. 755, no 2, 179-187 p.
Keyword [en]
Immunochemical detection, Detection (LC), Hollow fibres, Membranes, Biotin
National Category
Analytical Chemistry
Identifiers
URN: urn:nbn:se:hh:diva-24129DOI: 10.1016/S0021-9673(96)00620-6ISI: A1996WA19300003PubMedID: 8997748Scopus ID: 2-s2.0-0030572984OAI: oai:DiVA.org:hh-24129DiVA: diva2:679526
Note

This work was financially supported by the EC-project EC No EV5V-CT92-0109. Gilson Medical Electronics, Villiers-le-Bel, France, is acknowledged for instrumental support. The gift of hollow fibres from X-Flow BV, Almelo, Netherlands, is appreciated.

Available from: 2013-12-16 Created: 2013-12-09 Last updated: 2015-03-06Bibliographically approved

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