Laminin Biotin (mouse tumor)

AT

1-2 Weeks

15

Laminin-1 is purified from EHS tumor tissue and is free of the laminin binding protein entactin which is a common contaminant in some laminin preparations (150 kDa). Protein purity is determined by scanning densitometry of Coomassie Blue stained protein on a 4-20% polyacrylamide gel. The laminin is >90% pure (Figure 1).
The protein is modified to contain covalently linked biotins at random surface lysines. An activated ester of biotin is used to label the protein. The biotin label has been modified to contain a 14 atom spacer between the biotin and the laminin, this improves the accessibility of biotin to avidin/streptavidin.
Labeling efficiency is determined by the ability to detect 10 ng of biotinylated laminin using alkaline phosphatase labeled streptavidin (Figure 2). Laminin runs as individual subunits on SDS-PAGE with an apparent molecular weight of 400 and 225 kDa (Figure 1). LMN03 is supplied as an white lyophilized powder. Each vial of LMN03 contains 20 μg protein.

Serial dilutions of biotinylated laminin were separated by electrophoresis on a 4-20% SDS-polyacrylamide gel,  blotted to a PVDF membrane, probed with a 1:500 dilution of strepavidin alkaline phosphatase (Sigma) and detected with 1-Step NBY/BCIP reagent (Pierce). Lane 1, 100 ng; Lane 2,  50 ng; Lane 3, 40 ng; Lane 4, 20 ng; Lane 5, 10 ng of biotinylated laminin. Lane M, SeeBlue molecular weight markers (Invitrogen). Arrows indicate biotinylated laminin subunits (400kD and 225 kD).