False positive immunostaining of Locusta neurosecretory cells with a variety of preimmune sera.
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.
Tips
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A
Zoological Institute, Katholieke Universiteit Leuven, Belgium.
Schoofs
L
L
Paemen
L
L
Hendrickx
K
K
De Loof
A
A
D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D001923:Brain Chemistry / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D005189:False Positive Reactions; D005260:Female; D006110:Grasshoppers / Q000378:metabolism*; D006168:Guinea Pigs; D007106:Immune Sera / Q000276:immunology*; D015151:Immunoblotting; D007136:Immunoglobulins / Q000276:immunology / Q000302:isolation & purification; D007150:Immunohistochemistry; D050356:Lipid Metabolism; D008297:Male; D051379:Mice; D009490:Neurosecretory Systems / Q000166:cytology / Q000378:metabolism*; D010544:Peroxidases / Q000378:metabolism; D014176:Protein Biosynthesis; D011817:Rabbits; D013154:Spleen / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D016707:Tissue Fixation
D000906:Antibodies; D007106:Immune Sera; D007136:Immunoglobulins; D010544:Peroxidases