Ibarrola I, Sanz M L, Gamboa P M, Mir A, Benahmed D, Ferrer A, Arilla M C, Martínez A, Asturias J A
Departamento de Investigación y Desarrollo, Bial-Arístegui, Bilbao, Spain.
Clin Exp Allergy. 2004 Feb;34(2):303-9. doi: 10.1111/j.1365-2222.2004.01859.x.
Allergoids are widely used in specific immunotherapy (SIT) for the treatment of IgE-mediated allergic diseases, but all techniques for standardization of conventional allergic extracts may not be appropriate for standardization of a glutaraldehyde (GA)-modified extract because of the unique characteristics of these extracts.
To assess an accurate methodology for standardization of chemically modified extracts.
GA-modified extracts from Parietaria judaica pollen were purified by diafiltration. Biochemical properties were investigated by determination of amino groups, chromatography, and SDS-PAGE. The IgE-binding activity was determined by skin prick test, enzyme allergosorbent test inhibition, basophil activation, and histamine release tests. Peripheral blood mononuclear cells (PBMCs) from P. judaica pollen-allergic subjects were stimulated with either native or allergoid extracts, and proliferation was measured.
Biochemical data indicated a high degree of allergen polymerization resulting in extract components higher than 100 kDa. IgE-binding activity, both in vivo and in vitro, was reduced by more than 99.8%. Both allergen and allergoid induced PBMC proliferation and synthesis of blocking IgG antibodies at similar rates. Moreover, no evidence of introduction of new determinants by chemical modification was found.
The preparation of GA-modified extracts by diafiltration is faster and more reliable than previous chromatographic methods. These modified extracts have drastically reduced their allergenicity while maintaining their immunogenicity, and therefore they can be used in safer and shortened schedules of SIT.
变应原类制剂广泛应用于特异性免疫疗法（SIT）治疗IgE介导的过敏性疾病，但由于这些提取物的独特特性，常规变应原提取物的所有标准化技术可能不适用于戊二醛（GA）修饰提取物的标准化。
评估一种用于化学修饰提取物标准化的准确方法。
通过渗滤法纯化来自墙草花粉的GA修饰提取物。通过测定氨基、色谱法和SDS-PAGE研究其生化特性。通过皮肤点刺试验、酶变应原吸附试验抑制、嗜碱性粒细胞活化和组胺释放试验测定IgE结合活性。用天然或变应原类提取物刺激来自墙草花粉过敏受试者的外周血单个核细胞（PBMC），并测量其增殖情况。
生化数据表明变应原高度聚合，导致提取物成分高于100 kDa。体内和体外的IgE结合活性均降低了99.8%以上。变应原和变应原类制剂以相似的速率诱导PBMC增殖和封闭性IgG抗体的合成。此外，未发现化学修饰引入新决定簇的证据。
通过渗滤法制备GA修饰提取物比以前的色谱方法更快、更可靠。这些修饰提取物的致敏性大幅降低，同时保持其免疫原性，因此可用于更安全、更短疗程的SIT。
