The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use 0.5-1µg for 106 cells. ab170191-Mouse monoclonal IgG2a, is suitable for use as an isotype control with this antibody.
Use a concentration of 1 - 5 µg/ml. Predicted molecular weight: 48 kDa.
Use at an assay dependent concentration. Detection limit is approximately 3 ng/ml as a capture antibody when used against the immunogen.
May have a regulatory role in the membrane interactions during trafficking of synaptic vesicles at the active zone of the synapse. It binds acidic phospholipids with a specificity that requires the presence of both an acidic head group and a diacyl backbone. A Ca(2+)-dependent interaction between synaptotagmin and putative receptors for activated protein kinase C has also been reported. It can bind to at least three additional proteins in a Ca(2+)-independent manner; these are neurexins, syntaxin and AP2.
Belongs to the synaptotagmin family. Contains 2 C2 domains.
The first C2 domain mediates Ca(2+)-dependent phospholipid binding. The second C2 domain mediates interaction with SV2A and STN2.
Anti-Synaptotagmin 1 antibody (ab77314) at 5 µg/ml + immunogen at 0.2 µg
Secondary Goat Anti-Mouse IgG (H&L)-HRP Conjugate at 1/5000 dilution Developed using the ECL technique
Predicted band size : 48 kDa Observed band size : 65 kDa (why is the actual band size different from the predicted?) The observed band size may not correspond to the predicted protein molecular weight as the immunogen (recombinant tagged protein) was used as the sample. Molecular weight of the tag alone is 26 kDa.
Western blot - Synaptotagmin 1 antibody (ab77314)
Anti-Synaptotagmin 1 antibody (ab77314) at 5 µg/ml + Jurkat cell lysate at 25 µg
Secondary Goat Anti-Mouse IgG (H&L)-HRP Conjugate at 1/2500 dilution Developed using the ECL technique
Overlay histogram showing SH-SY5Y cells stained with ab77314 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab77314, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2a [ICIGG2A] (ab91361, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.