Isolation of Der pI, the Dermatophagoides pteronyssinus major mite allergen, from a crude mite culture extract, purification by ion-chromatography, and comparison between the material obtained and a cDNA-coded Der pI

• Published in: Journal of Chromatography A 1992-05, Vol.599 (1), p.105-111
• Main Authors: Dandeu, J.-P., Rabillon, J., Lux, M., David, B., Guillaume, J.-L., Camoin, L.
• Format: Article
• Language: English
• Subjects: allergens; Antigens, allergens; Biological and medical sciences; Dermatophagoides pteronyssinus; extracts; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Immediate hypersensitivity. Allergy. Anaphylaxis, etc; Immunobiology; ion exchange chromatography; purification
• ISSN: 0021-9673
• DOI: 10.1016/0021-9673(92)85462-3

A high degree of purity is a prerequisite for an allergen prepration to be suitable for clinical diagnosis and therapy. A pure allergen can easily be obtained from a crude extact by using an immunosorbent prepared with highly specific monoclonal antibodies or from a cDNA-coded material. However, up to now none of these methods has been performed on a process scale. Here large-scale purification is defined as a process in which a crude Dermatophagoides pteronyssinus mite culture extract is essentially fractionated by acetone and ammonium sulphate precipitations followed by anion-exchange high-performance liquid chromatography. A high yield of a very pure Der pI allergen is obtained during the first isocratic run, as shown by sodium dodecylsulphate-polyacrylamide gel electrophoresis, capillary electrophoresis, chromatofocusing and a two site monoclonal antibody enzyme-linked immunosorbent assay. Microsequencing revealed that the 25-residue sequence obtained is entirely in agreement with the sequence derived from the cDNA of Der pI.