Anti-Synaptotagmin antibody [ASV30]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Synaptotagmin antibody [ASV30] (AB13259)

ICC/IF image of ab13259 stained PC12 cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab13259, 5μg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43μM.

This image was generated using the ascites version of the product.

• IP

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Immunoprecipitation - Anti-Synaptotagmin antibody [ASV30] (AB13259)

Synaptotagmin was immunoprecipitated using 0.5mg Mouse Brain tissue lysate, 5μg of Mouse monoclonal to Synaptotagmin and 50μl of protein G magnetic beads (+). No antibody was added to the control (-).
The antibody was incubated under agitation with Protein G beads for 10min, Mouse Brain tissue lysate lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
Proteins were eluted by addition of 40μl SDS loading buffer and incubated for 10min at 70^oC; 10μl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab13259.
Secondary : Goat polyclonal to mouse IgG light chain specific (HRP) at 1/20,000 dilution.
Band : 64kDa; Synaptotagmin

This image was generated using the ascites version of the product.

All lanes:

Immunoprecipitation - Anti-Synaptotagmin antibody [ASV30] (ab13259)

Predicted band size: 47 kDa

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