AMBP | Bikunin (50 ĩl)
AS06 173 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Human
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human recombinant bikunin, UniProt: P02760, cleavage product no. PRO_0000017887
free bikunin 60-80 kDa
Panel A: Coomasie stained gel. Panel B: 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) 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.
Bikunin is a Kunitz-type protease inhibitor synthesized in the liver. The AMBP gene resides in the lipocalin cluster and encodes alpha-1-microglobulin together with a Kunitz-type proteinase inhibitor, bikunin. The gene is translated into the alpha-1-microglobulin-bikunin precursor, which is subsequently cleaved and the two proteins are secreted to the blood separately. Bikunin is found in blood and tissues both in free form and associated with heavy chains forming the inter-alpha-inhibitor family of proteins. Bikunin is a protease inhibitor, a component of extracellular matrix and exhibits anti-inflammatory activity in protection against cancer and inflammation.
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