A simple, rapid, and sensitive scintillation proximity assay for the determination of levels of guinea-pig interleukin-5 in bronchoalveolar lavage samples.

Abstract

This article describes the development and validation of a scintillation proximity assay (SPA) sensitive for guinea-pig interleukin-5 (IL-5). SPA beads were coated with TRFK-5, a monoclonal antibody directed against mouse IL-5, which is known also to bind guinea-pig IL-5. The assay is a simple competitive binding assay between [125I]-rh-IL-5 and the IL-5, in a sample of guinea-pig bronchoalveolar lavage fluid (BALF), for the binding site on the TRFK-5-coated beads. IL-5 levels in BALF ([IL-5]BALF) were shown to increase in guinea-pigs sensitized to ovalbumin (OvA) and challenged with an OvA inhalation. This occurred at a time (24 h) after challenge when there was also a marked eosinophilia. The assay was validated by treating guinea-pigs with a second antibody, Genzyme 2374-01, directed against IL-5. Treatment with this antibody resulted in a significant reduction of the antigen-induced eosinophilia and concentration of [IL-5]BALF. This observation confirms that the IL-5 identified in BALF also cross-reacts with the antibody Genzyme 2374-01. Interestingly, plasma from sensitized, but unchallenged, guinea-pigs also contained detectable levels of IL-5, and the stimulation of plasma protein extravasation (PPE) within the airways with inhaled histamine also induced a rise in [IL-5]BALF. These observations suggest that the plasma may be an additional source of the IL-5 present in the airways of antigen-challenged guinea-pigs.

• Authors: H Danahay, K J Broadley, P J McCabe, A T Nials, S Sanjar
• Journal: Journal of pharmacological and toxicological methods (J Pharmacol Toxicol Methods) Vol. 37 Issue 3 Pg. 161-6 (Apr 1997) ISSN: 1056-8719 [Print] United States
• PMID: 9253752 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antibodies, Monoclonal
• Blood Proteins
• Interleukin-5
• Recombinant Proteins
• Histamine
• Ovalbumin

Topics

• Administration, Inhalation
• Animals
• Antibodies, Monoclonal (metabolism, pharmacology, therapeutic use)
• Antibody Formation
• Binding, Competitive
• Blood Proteins (metabolism)
• Bronchoalveolar Lavage Fluid (chemistry, cytology)
• Eosinophilia (chemically induced, drug therapy)
• Guinea Pigs
• Histamine (administration & dosage, immunology, toxicity)
• Interleukin-5 (analysis, immunology, metabolism)
• Male
• Mice
• Microspheres
• Ovalbumin (administration & dosage, immunology, toxicity)
• Rats
• Recombinant Proteins (analysis, metabolism)
• Scintillation Counting