Tabar A I, Sanz M L, Oehling A
Departamento de Alergología, Facultad de Medicina, Universidad de Navarra, Pamplona, Spain.
Allergol Immunopathol (Madr). 1990 Jan-Feb;18(1):13-7.
In a series of works done on humans patterned after animal experimental models, we created an "in vitro" model which allowed us to produce soluble IgE regulator factors. We observed its biological activity in IgE synthesis and constructed changes that protein fractions from affinity chromatography (fractions enriched in soluble regulator IgE factors) exercised on IgE receptor expression. We then decided to determine the molecular weight of each one of the active protein fractions. We performed electrophoresis on the gel of polyacrylamide (Weber-Osborn technique). Protein bands were processed, with the aim of isolating the gel and quantifying the biological activity. The protein fractions obtained from the chromatographies with affinity to CON-A sepharose were subjected to electrophoresis in polyacrylamide gel, with the aim of obtaining molecular weights of proteins comprising it. Using rosette technique, there were no changes seen in Fc epsilon expression just as what happened in the experiment with material fractionated by chromatography. However, these changes are produced and are statistically significant using the immunofluorescence technique. The highest inhibition of rosettes or immunofluorescence is that of the protein band of 10,000 daltons extracted in NaCl and that of 20,000 daltons extracted in PBS. Both effects are similar. The inhibition reached by the 20,000 dalton band in NaCl is less although there are no significant differences. The material extracted in PBS of the band between 6.9 and 7.2 mm of gel (G4) did not have any changes. None of the proteins added to the culture medium had any effects of receptor gamma expression. The average values of the receptor expression was similar to basal levels.
在一系列以动物实验模型为蓝本开展的人体研究中，我们创建了一个“体外”模型，该模型使我们能够产生可溶性IgE调节因子。我们观察了其在IgE合成中的生物活性，并构建了亲和层析得到的蛋白组分（富含可溶性调节IgE因子的组分）对IgE受体表达的影响。然后，我们决定测定每个活性蛋白组分的分子量。我们采用聚丙烯酰胺凝胶电泳（韦伯-奥斯本技术）。对蛋白条带进行处理，目的是分离凝胶并定量生物活性。对与伴刀豆球蛋白A琼脂糖有亲和力的层析所得蛋白组分进行聚丙烯酰胺凝胶电泳，以获得其所含蛋白质的分子量。使用玫瑰花环技术，Fcε表达未见变化，就如同层析分离的物质实验中所发生的那样。然而，使用免疫荧光技术时会产生这些变化，且具有统计学意义。对玫瑰花环或免疫荧光的最高抑制作用是在NaCl中提取的10,000道尔顿蛋白条带以及在PBS中提取的20,000道尔顿蛋白条带。两种效应相似。虽然没有显著差异，但在NaCl中20,000道尔顿条带的抑制作用较小。在凝胶6.9至7.2毫米之间（G4）的条带中，PBS提取的物质没有任何变化。添加到培养基中的任何蛋白质对γ受体表达均无影响。受体表达的平均值与基础水平相似。
