GP | glycogen phosphorylase
AS09 455 | clonality:polyclonal | host:rabbit | reactivity:chicken, human, mouse, rabbit, rat, T.vaginalis G3
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1: 2000 (WB), 5 µg (IP)
|Expected | apparent MW||
human, mouse, rabbit,rat, Trichomonas vaginalis G3
bovine, dog, hen,horse,pig, Xenopus laevis, Zebrafish, atlantic salmon, Drosophila melanogaster
|Not reactive in||
no confirmed exceptions from predicted reactivity known in the moment
samples used to test this antibody were: muscle and liver homogenates, purified glycogen. This antiboy will recogize glycogen phosphorylase from both liver and muscle. It has been used in ICC on human primary myotubes
|Selected references||Dandanell et al. (2016). Maintaining a clinical weight loss after intensive lifestyle intervention is the key to cardiometabolic health. pii: S1871-403X(16)30107-7. doi: 10.1016/j.orcp.2016.09.009.
Bowker and Zhuang (2015). Relationship between water-holding capacity and protein denaturation in broiler breast meat. Poult Sci. 2015 May 25. pii: pev120.
Zhu et al. (2011). High post-mortem temperature combined with rapid glycolysis induces phosphorylase denaturation and produces pale and exudative characteristics in broiler Pectoralis major muscles. Meat Scienc
(1) rat liver homogenate (50 µg of total protein), (2) 10 ug purified rat liver glycogen (see Parker et al, BBRC 2007), (3) Human skeletal muscle homogenate (50 µg of total protein), (4) GP IP from 500 ug Human skeletal muscle homogenate were resuspended into SDS PAGE sample buffer, boiled, electrophoresed on a pre-cast 4-15% gradient gel (Invitrogen, CA, USA) and transferred to PVDF membrane. Following blocking in 5% milk / PBST, membranes were probed with GP antibodies (diluted 1/1000) for 1h at room temperature with rocking, followed by washing in 3 times for 5 min in PBS-T at room temperature with agitation. Blots were incubated in secondary antibody (Protein-G coupled to horse radish peroxidase conjugated – Bio-Rad) diluted 1:3000 for 1h at room temperature with agitation. The blots were washed as above and developed for 5 min with ECL (Invitrogen, CA, USA). Images were detected using the Fuji LAS-3000 system.
To obtain GP-bound immunoprecipitates, 5 ug GP antibody was incubated with 500 ug human skeletal muscle homogenate together with 1 ml IP wash buffer (50 mM Hepes pH 7.5, 150 mM NaCl, 10% glycerol, 1% NP-40, 1 mM EGTA, 10 mM NaPPi and 100 mM NaF) and incubated at 4°C with mixing for 90 minutes. 100 ul, 20% Protein-A Sepharose was then added for a further 30 minutes at 4°C with mixing. Immunoprecipitates were collected by centrifugation at 7,000 rpm for 1 minutes and washed three times with IP wash buffer followed by resuspension into SDS PAGE sample buffer, boiling and electrophoresis as above.
The GP antibody did not immunoprecipitate glycogen phosphorylase from rat liver.
Courtesy of Dr. Melissa Stuart, Microbiology/Immunology Kirksville College of Osteopathic Medicine, USA
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