Molecular Structure and Interaction of Recombinant Human Type XVI Collagen

Collagen XVI is a neural component of the developing and regenerating dorsal root ganglia extracellular matrix

“…Western blots were performed as previously described (Kassner et al, 2004) on dissociated embryonic neurons conditioned media after 48 h of culture. Control experiments were done on neuronal culture medium and conditioned medium from human dermal fibroblasts.…”

“…The following primary antibodies were used: guinea-pig anticollagen XVI antibody (1/75), (Kassner et al, 2004), rabbit anti-tau or mouse monoclonal anti-NF 160 antibody from Sigma (1/500), rabbit anti-fibronectin (1/100). NeuroTrace from Invitrogen (1/200) was used for vizualisation of cellular morphology.…”

Collagen XVI induces formation of focal contacts on intestinal myofibroblasts isolated from the normal and inflamed intestinal tract

“…Notably, as in the DEJ fibrillin-1 and collagen XVI are partly but not always co-localized. This could be explained by our previous findings, that recombinant fibrillin-1 fragments interact strongly with collagen XVI, however under physiological salt concentrations with lower affinity (Kassner et al, 2004). Preliminary data reveal no association of collagen XVI to collagen I fibrils as shown previously also for the dermis (Kassner et al, 2003) which leaves fibrillin-1 as a putative binding partner for collagen XVI in the bowel wall.…”

“…Blocking was performed for 1 h at RT with 5% dry milk in T-PBS. The blots were longitudinally cut above the 50 kDa standard and the upper part was incubated with protein-A purified Col XVI -GP antibody (Kassner et al, 2004) overnight at 4°C (dilution 1:1000) and the lower part with a mouse ß-actin antibody (ab21250-100, Abcam). Horseradish peroxidase conjugated secondary antibody was incubated for 1 h at RT (1:30,000 dilution; Sigma, USA).…”

“…96-well plates (MaxiSorb, Falcon) were coated with poly-L-lysine (5 µg/ml; Sigma, USA), collagen I (10 µg/ml; Sigma USA) and recombinant collagen XVI (Kassner et al, 2004) (10 µg/ml and 30 µg/ml) overnight at 4°C, followed by two washing steps with PBS and an additional coating of unspecific binding sites with 1% BSA (w/v) (Biomol, Germany) solution for 2 h at RT. The plates were washed two times with PBS and stored at 4°C until usage.…”