Characterization of small-field bistratified amacrine cells in macaque retina labeled by antibodies against synaptotagmin-2.

Macaque retinae were immunostained with monoclonal antibodies directed against the protein synaptotagmin-2 (Syt2). Syt2 was localized in a population of small-field amacrine cells, whose cell bodies formed a regular mosaic within the inner nuclear layer, indicating they represent a single amacrine cell type. The labeled amacrine cells had a bistratified appearance with a dense dendritic plexus in the OFF-layer and only a few lobular processes extending into the ON-layer of the inner plexiform layer, similar to A8 amacrine cells described in cat and human retina. Syt2-labeled cells were immunoreactive for glycine but lacked immunoreactivity for γ-aminobutyric acid (GABA), suggesting they use glycine as their neurotransmitter. The density of these cells increases from ∼200/mm(2) in peripheral retina to ∼1,400/mm(2) in central retina. Their bipolar cell input was studied by immunolabeling experiments using various bipolar cell markers combined with CtBP2, a marker of presynaptic ribbons. Our data show that Syt2-labeled amacrine cells receive input from both OFF and ON cone bipolar cells, as well as from rod bipolar cells. The OFF input is dominated by the diffuse bipolar cell DB1 (44%) and the OFF midget bipolar cell (38%). Here we describe a population of bistratified small-field amacrine cells closely resembling A8 amacrine cells and their cone-dominated bipolar cell input. J. Comp. Neurol. 521:709-724, 2013. © 2012 Wiley Periodicals, Inc.

Synaptotagmin II peptide-bead conjugate for botulinum toxin enrichment and detection in microchannels.

This paper reports an enrichment platform for botulinum neurotoxin type B (BoNT/B) that has been realized through the fusion of bioconjugation chemistry and microfluidics. Micrometer-sized magnetic beads were conjugated to a 22mer synthetic peptide derived from the synaptotagmin II (Syt II) neuronal protein that is specific for BoNT/B binding. Exposure to BoNT/B in buffer, whole milk and fruit juices resulted in toxin capture, which was confirmed using immunofluorescence. Peptide-modified beads were integrated into arrayed, polymeric microfluidic channels, and all assay steps, from capture to detection, were performed directly in the microchannels, thereby simplifying assay utility and increasing throughput relative to existing detection methodologies. Our sensitive microscale approach required only 7 µL of intentionally adulterated sample without any pre-processing (i.e. dilution, centrifugation, filtering), and with a "hands-on" time of only 1 h to detect 16.6 pg of BoNT/B in whole milk.