Chickens were immunized with ECS, Enterokinase Cleavage Site (xxxDDDDK) conjugated to KLH. Antibody was isolated by affinity chromatography using the peptide immobilized on solid support. Antibody concentration was determined by extinction coefficient: absorbance at 280 nm of 1.4 equals 1.0 mg of IgG. By ELISA and Western blot, this antibody reacts with FLAGTM.
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Detection of ECS (DDDK)-tagged Protein by western blot. Samples: 200, 100, or 50 ng of E. coli whole cell lysate expressing a multi-tag fusion protein. Antibodies: Affinity purified, chicken anti-ECS (DDDK) antibody A190-200A used for WB at 0.2 µg/ml (1:5,000). Detection: Chemiluminescence with an exposure time of 30 seconds.