Characterization of the syntaxin 2 and 3 anti-sera. (A) Equal amounts of syntaxin 2, 3, and 4 cytosolic domain GST fusion proteins (2 μg) were incubated with thrombin (0.3 U) for two hours to cleave off the GST (27 kDa, indicated with arrow) and analysed by SDS-PAGE, transferred to nitrocellulose filters and probed with syntaxin 2 or 3 anti-serum followed by alkaline phosphatase-conjugated secondary antibodies. (B)The Western blotting of enriched membrane fraction of NRK cells were probed with syntaxin 2 anti-serum (lane 1), syntaxin 2 anti-serum pre-incubated with syntaxin 2-GST protein (lane 2), syntaxin 3 anti-serum (lane 3) and syntaxin3 anti-serum preincubated with syntaxin 3-GST protein (lane 4) as well as horseradish peroxidase conjugated secondary antibodies. Each lane contains 25 μg protein. All the samples were boiled for 3 minutes in the presence of 2% SDS in Laemmli sample buffer.
Band and Kuismanen BMC Cell Biology 2005 6:26 doi:10.1186/1471-2121-6-26