FALSE-POSITIVE IMMUNOSTAINING OF LOCUSTA NEUROSECRETORY-CELLS WITH A VARIETY OF PREIMMUNE SERA

A large number of antisera directed against vertebrate neuropeptides h ave been reported to yield positive staining when applied to insect br ains. In most cases, the preimmune serum of the same animal in which t he antiserum was developed is not available for testing in control exp eriments. We have experienced that a large percentage of preimmune ser a, as well as a culture medium for hybridomas, stain cell populations and fibers in the central nervous system of the insect Locusta migrato ria. 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 extract s indicates the presence of two immunoreactive 27-kDa bands. These ban ds could also be visualized in other tissue extracts such as muscle, m idgut, Malpighian tubules, and fat body of Locusta. The brain of other insect species, such as Periplaneta americana, Leucophaea maderae, an d Neobellieria bullata were devoid of the false immunopositive reactio n. There is no easy way to eliminate this type of immunoreaction. It f ollows 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 se ll or exchange sera together with their corresponding preimmune sera. (C) 1997 Academic Press.