Tips A, Schoofs L, Paemen L, Hendrickx K, De Loof A
Zoological Institute, Katholieke Universiteit Leuven, Belgium.
Gen Comp Endocrinol. 1997 May;106(2):231-40. doi: 10.1006/gcen.1996.6869.
A large number of antisera directed against vertebrate neuropeptides have been reported to yield positive staining when applied to insect brains. In most cases, the preimmune serum of the same animal in which the antiserum was developed is not available for testing in control experiments. We have experienced that a large percentage of preimmune sera, as well as a culture medium for hybridomas, stain cell populations and fibers in the central nervous system of the insect Locusta migratoria. Purification of these preimmune sera on a Protein A and Protein G support indicates that the reaction is due to preexisting antibodies of the IgG class. Western analysis of brain and nervous tissue extracts indicates the presence of two immunoreactive 27-kDa bands. These bands could also be visualized in other tissue extracts such as muscle, midgut, Malpighian tubules, and fat body of Locusta. The brain of other insect species, such as Periplaneta americana, Leucophaea maderae, and Neobellieria bullata were devoid of the false immunopositive reaction. There is no easy way to eliminate this type of immunoreaction. It follows that when affinity chromatographic purification of the antibody is not feasible, it is essential to include in the control procedure, the preimmune serum of the animal that was used for the production of the antiserum. This means that it should become common practice to sell or exchange sera together with their corresponding preimmune sera.
据报道，大量针对脊椎动物神经肽的抗血清应用于昆虫脑时会产生阳性染色。在大多数情况下，用于制备抗血清的同一动物的免疫前血清无法用于对照实验检测。我们发现，很大比例的免疫前血清以及杂交瘤细胞的培养基，会对蝗虫中枢神经系统中的细胞群体和纤维产生染色。在蛋白A和蛋白G载体上对这些免疫前血清进行纯化表明，该反应是由预先存在的IgG类抗体引起的。对脑和神经组织提取物进行的蛋白质印迹分析表明存在两条免疫反应性的27 kDa条带。这些条带在其他组织提取物中也可见，如蝗虫的肌肉、中肠、马氏管和脂肪体。其他昆虫物种，如美洲大蠊、马德拉蜚蠊和大绿蝇的脑则没有这种假免疫阳性反应。没有简单的方法可以消除这种类型的免疫反应。因此，当抗体的亲和层析纯化不可行时，在对照程序中加入用于制备抗血清的动物的免疫前血清至关重要。这意味着出售或交换血清及其相应的免疫前血清应该成为一种惯例。
