Mango allergen components analysis by three different extracting methods

Objective To evaluate three extracting methods for mango proteins and analyze the protein components which could bind with serum specific IgE( sIgE) of allergy patients. To provide an optimal antigen extraction protocol for clinical detection of allergen specific IgE. Methods Proteins were extracted from mango using trichloroacetic acid precipitation method,trichloroacetic acid /acetone precipitation method and lysate extraction method,respectively. The total protein components were analyzed by SDS-PAGE and two-dimensional electrophoresis. Immunoblot technique and two-dimensional immunoblot technique were used to identify protein components which could combine with specific IgE. Results Protein components extracted by three methods were different. The single patient serum showed different affinity and binding sites in the immunoblot assay. Protein components from trichloroacetic acid precipitation method and trichloroacetic acid /acetone precipitation method reacted with patient serum mainly at 30,40,44,47 and 90 kD,while that from lysate extraction method reacted with patient serum at mainly 23,32,40,46,73 and 90 kD. Conclusion Three kinds of extraction methods had different characteristics,and trichloroacetic acid precipitation method was more complete,reproducible,easy to operate and could meet the requirements of mango sIgE detection.