The specificity of nc82 against BRP protein has been demonstrated by: 1) the expression pattern of GFP-tagged bruchpilot driven under tissue-specific drivers, which matches nc82 signals in wing discs and tracheal cells, and is also targeted to the active zone of larval NMJ boutons (Wagh et al., 2006 (link)); 2) western blots of adult head extracts using nc82 (Wagh et al., 2006 (link)); and 3) the loss of immune-expression in brp mutant neuromuscular junctions and rescue by expression of BRP in brp mutants (Kittel et al., 2006 (link)). nc82 has been widely used to label synaptic sites in Drosophila, based largely on the pattern of labeling demonstrated at neuromuscular junctions and fly photoreceptor synapses (Hamanaka and Meinertzhagen, 2010 (link)), but reports of its specificity are mostly not complete for synapses of the CNS. In particular, nc82 labels the platform of the T-bar ribbon, and not only has nc82 not been shown to label the platforms at CNS synapses, but not all synapses in the CNS have such organelles (Butcher et al., 2012 (link)).
The specificities of the three epitope-tagged antibodies, rat anti-FLAG, rabbit anti-hemagglutinin (anti-HA), and mouse anti-V5, are validated by the internal controls of the flip-out approach in that: 1) expression patterns differ by GAL4 line and 2) the extent of labeling varies from no label to dense label even though the GAL4 drivers are reasonably broad.
The specificities of the three epitope-tagged antibodies, rat anti-FLAG, rabbit anti-hemagglutinin (anti-HA), and mouse anti-V5, are validated by the internal controls of the flip-out approach in that: 1) expression patterns differ by GAL4 line and 2) the extent of labeling varies from no label to dense label even though the GAL4 drivers are reasonably broad.
