Chen Z, Van Kampen V, Raulf-Heimsoth M, Baur X
Research Institute for Occupational Medicine (BGFA), Ruhr-University of Bochum, Germany.
Clin Exp Allergy. 1996 Apr;26(4):406-15.
The rubber elongation factor in Hevea rubber (Hev b 1) is one of the most important latex allergen and is leading cause of latex type 1 hypersensitivity in children with spina bifida.
The aim of this study was to define the allergenic and antigenic epitopes of Hev b 1.
The immunoglobulin- (Ig)E and IgG antibody binding sites on Hev b 1 allergen were delineated by enzyme linked immunosorbent assay (ELISA) using synthetic overlapping peptides covering the whole Hev b 1 sequence. In order to improve the binding capacity and specificity all peptides were biotinylated at the N-terminal end via a 6-aminohexanoic acid as spacer and then adsorbed to streptavidin pre-coated microtitre plates. Fine mapping to define the essential amino acid residues for the antibody binding was achieved by using overlapping peptides with one amino acid offset.
It was demonstrated that the IgE epitopes were located in different regions of Hev b 1 including the C-terminal segment (121-137) and the segments with amino acid residues of 30-49 and 46-64. Two monoclonal antibodies (MoAbs) II2F3 and II4G9 raised against purified Hev b 1 recognized the C-terminal segment only. The results of epitope mapping with three rabbit antisera revealed that five positive peptides, including the epitope peptides 31-49, 46-64 and 121-137, were involved in the antibody-binding sites. Fine mapping on the segments 46-64 and 121-137 showed that the two MoAbs reacted with the peptide 125-134 in the C-terminal region, whereas the peptide with amino acids 124-134 was essential for recognition by human IgE antibodies. Epitopes to rabbit polyclonal IgG and human IgE were also found to be involved in the amino acid residues of 47-59.
Our results indicate that the most allergenic/antigenic portions of Hev b 1 allergen are the C-terminal region and the region with amino acid residues of 31-64. In both regions, the minimal IgE-binding epitope is almost identical with the IgG-binding epitope.
巴西橡胶树中的橡胶延伸因子（Hev b 1）是最重要的乳胶过敏原之一，是脊柱裂患儿乳胶Ⅰ型超敏反应的主要原因。
本研究旨在确定Hev b 1的致敏和抗原表位。
使用覆盖整个Hev b 1序列的合成重叠肽，通过酶联免疫吸附测定（ELISA）描绘Hev b 1过敏原上的免疫球蛋白（Ig）E和IgG抗体结合位点。为了提高结合能力和特异性，所有肽在N末端通过6-氨基己酸作为间隔臂进行生物素化，然后吸附到预包被链霉亲和素的微量滴定板上。通过使用一个氨基酸偏移的重叠肽进行精细定位，以确定抗体结合的必需氨基酸残基。
结果表明，IgE表位位于Hev b 1的不同区域，包括C末端片段（121-137）以及氨基酸残基为30-49和46-64的片段。两种针对纯化的Hev b 1产生的单克隆抗体（MoAbs）II2F3和II4G9仅识别C末端片段。用三种兔抗血清进行表位定位的结果显示，五个阳性肽，包括表位肽31-49、46-64和121-137，参与了抗体结合位点。对片段46-64和121-137的精细定位表明，这两种单克隆抗体与C末端区域的肽125-134反应，而氨基酸为124-134的肽对于人IgE抗体的识别至关重要。还发现兔多克隆IgG和人IgE的表位涉及47-59的氨基酸残基。
我们的结果表明，Hev b 1过敏原的最具致敏性/抗原性的部分是C末端区域和氨基酸残基为31-64的区域。在这两个区域中，最小的IgE结合表位与IgG结合表位几乎相同。
