کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
5516117 | 1542310 | 2017 | 6 صفحه PDF | دانلود رایگان |
- IAC column was prepared for Ara h 2 isolation.
- The purification cost less time.
- Both Ara h 2 and its derivative could be separated by IAC.
Ara h 2 is considered a major allergen in peanut. Due to the difficulty of separation, Ara h 2 had not been fully studied. Immunoaffinity chromatography (IAC) column can separate target protein with high selectivity, which made it possible to purify Ara h 2 from different samples. In this study, IAC method was developed to purify Ara h 2 and its effect was evaluated. By coupling polyclonal antibody (pAb) on CNBr-activated Sepharose 4B, the column for specific extraction was constructed. The coupling efficiency of the IAC column was higher than 90%, which made the capacity of column reached 0.56Â mg per 0.15Â g medium (dry weight). The recovery of Ara h 2 ranged from 93% to 100% for different concentrations of pure Ara h 2 solutions in 15Â min. After using a column 10 times, about 88% of the column capacity remained. When applied to extract Ara h 2 from raw peanut protein extract and boiled peanut protein extract, the IAC column could recovery 94% and 88% target protein from the mixture. SDS-PAGE and Western blotting analysis confirmed the purified protein was Ara h 2, its purity reached about 90%. Significantly, the IAC column could capture dimer of Ara h 2, which made it feasible to prepared derivative of protein after processing.
Journal: Protein Expression and Purification - Volume 131, March 2017, Pages 85-90