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AS12 2220 | Clonality: Polyclonal | Host: Rabbit | Reactivity:Human influenza hemagglutinin (HA) tagged proteins (strong cross reactivity between 25 -20 kDa in Arabidopsis thaliana wildtype)
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KLH-conjugated synthetic peptide: YPYDVPDYA
depends on the MW of the protein which is HA-tagged
10 µg of total protein extracted freshly from mutant HEK293 cells with RIPA buffer and denatured at 65°C for 5 min were separated on 15% SDS-PAGE and blotted overnight to PVDF (pore size of 0.4 um), using wet transfer. Blot was blocked with 5% milk or 2 h at RT and then incubated in the primary antibody at a dilution of 1: 5 000 in a rotator for 2.5 h in 5% milk. The membrane was then placed in a clean container and rinsed 3 times for 5 min in TBS-T at RT with agitation. Blot was incubated in Agrisera matching secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, AS09 602) diluted to 1:20 000 in for 1 h at RT with agitation. The blot was washed as above and developed with Agrisera ECLSuperBright. Exposure time was 30 seconds.
Courtesy Dr. Małgorzata Wessels, Department of Medical Biochemistry and Biophysics, Umeå University, Sweden
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