Abstract
The Wuerzburg Hybridoma Library against the Drosophila brain represents a collection of around 200 monoclonal antibodies that bind to specific structures in the Drosophila brain. Here we describe the immunohistochemical staining patterns, the Western blot signals of one- and two-dimensional electrophoretic separation, and the mass spectrometric characterization of the target protein candidates recognized by the monoclonal antibodies aa2 and ab52 from the library. Analysis of a mutant of a candidate gene identified the Drosophila homolog of the Epidermal growth factor receptor Pathway Substrate clone 15 (Eps15) as the antigen for these two antibodies.
Highlights
The generation of hybridomas was one of the milestones in modern biology [1,2] leading to the production of monoclonal antibodies, one of the most important tools in biology
By the candidate approach the antigen of monoclonal antibodies (mAbs) nb33 which binds to pigment dispersing factor (PDF) containing neurons was identified as the PDF precursor protein [25]
In addition to the mAbs generated against brain homogenate our hybridoma library contains mAbs 3C11 and pok13 which were generated against bacterially expressed Drosophila synapsin and calbindin-32, respectively [30,31]
Summary
The generation of hybridomas was one of the milestones in modern biology [1,2] leading to the production of monoclonal antibodies (mAbs), one of the most important tools in biology. In line with our long standing research focus on synaptic proteins [26,27,28,29,30,31] we here describe the mAbs aa2 and ab52, their binding to synaptic neuropil of the adult brain, their immunohistochemical staining of the synaptic boutons of larval motor neurons, and the experiments leading to the identification of the homologue of Epidermal growth factor receptor Pathway Substrate clone 15 (Eps15) as the antigen they recognize in the Drosophila brain.
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