Preparation and activity identification of recombinant allergen rDer f 27 from Dermatophagoides farina
10.3969/j.issn.1000-484X.2024.10.025
- VernacularTitle:重组粉尘螨变应原rDer f 27制备及其活性鉴定
- Author:
Yaning REN
1
;
Dongmei ZHOU
;
Yuanfen LIAO
;
Ying ZHOU
;
Yubao CUI
;
Jie FEI
Author Information
1. 南京医科大学附属无锡人民医院,无锡 214023
- Keywords:
Dermatophagoides farinae;
Allergen;
Der f 27;
Immunoreactivity;
Immunogenicity;
Serine protease inhibitors
- From:
Chinese Journal of Immunology
2024;40(10):2168-2173
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To prepare recombinant protein of group 27 allergen of Dermatophagoides farinae(Der f 27),and to determine its immunoactivity.Methods:pET-28a(+)-Der f 27 plasmid was constructed and inserted into E.coli BL21(DE3)cells.After being expressed and purified,recombinant allergen rDer f 27 was obtained.IgE binding rates of rDer f 27 with sera from patients with allergic rhinitis induced by Dermatophagoides farinae was determined by ELISA and Western blot.PBMC from patients with allergic rhinitis and BESA-2B cells were cultured with rDer f 27 for 24 h,respectively,and cytokine expression was measured.Bioinformatics softwares were used to analyze physicochemical properties and structures of Der f 27.Results:pET-28a(+)-Der f 27 plasmids were prepared successfully and transformed into BL21(DE3)cells.After expressed and purified with PTG,SDS-PAGE and Western blot identified a band about 48 kD.IgE binding rates of rDer f 27 were 39.5%and 45.5%with sera from patients with allergic rhinitis allergic to Dermatophagoides farinae by IgE-ELISA and IgE-Western blot,respectively.Compared with control group,IL-6 and IL-8 expressions were increased in PBMC from patients with allergic rhinitis being cultured with rDer f 27(P<0.05);expressions of IL-10 and TGF-β were decreased in BESA-2B cells being cultured with rDer f 27,while IL-17A and IL-23A expressions were increased.Bioinformatics analysis showed that Der f 27 belong to serine protease inhibitor family and had universal structure and func-tion of this family.Secondary structure of Der f 27 was mainly composed of α-helix(42.62%)and random coil(35.60%).Conclusion:Recombinant allergen rDer f 27 has been prepared successfully with good immunoreactivity and immunogenicity,becoming one of important allergens of allergic rhinitis.
