AMBP | Bikunin (rat antibody)
AS04 039 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Rat
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|Recommended dilution||1 : 2000 (ELISA ),Assay dependent dilution (IHC), 1 : 2000 (WB)|
|Expected | apparent MW||
Free bikunin appears as a smear between 60-80 kDa, because the protein core carries sulfated proteoglycans.
The most common bikunin complexes are called pre-alpha-inhibitor (PaI) and inter-alpha-inhibitor (IaI) and appears as approximately 120 and 180 kDa
|Not reactive in||No confirmed exceptions from predicted reactivity are currently known.|
to be added when available
5 ug human (1), rat (2) or mouse (3) bikunin, expressed and purified from recombinant insect cells as described (ref 1), 8 µg of total protein from human (4), rat (5) or mouse (6) serum were mixed in sample buffer, containing 2% SDS and 1% mercaptoethanol, and denatured at 100°C for 2 min, and then separated on pre-cast 4-20 % gels bought from Bio-Rad (Mini-PROTEAN Tris-Glycine, Stain-Free, 4-20%) and blotted PVDF using semi-dry transfer. The membranes were blocked with 1% gelatin buffer 20 min at room temperature (RT) with agitation, and then incubated in the primary antibody at a dilution of 1: 2000 in 0.5% gelatin and 0.25% Tween-20 overnight at 4°C with agitation. The antibody solution was decanted and the membrane was rinsed 4x7 min with in 0.5% gelatin and 0.25% Tween-20, and incubated in secondary antibody (goat anti-rabbit IgG-Alexa 647, from Molecular Probes Inc) diluted to 10 ug/ml in 0.5% gelatin and 0.25% Tween-20 for 1h at RT with agitation. The blot was washed as above and developed using a ChemiDoc™ MP System (Bio-Rad) with the Alexa 650-settings.
Reference 1. Falkenberg et al. (2001). Expression of a functional proteinase inhibitor capable of accepting xylose: bikunin. Arch Biochem Biophys 387: 99-106
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