Combined effects of glutaraldehyde and riboflavin/uv365 on the self-assembly of type I collagen molecules observed with atomic force microscopy

Abstract

The aim of this study was to demonstrate the effect of different treatments on collagen self-assembly, the morphological and the qualitative analysis of collagen assembly were measured by atomic force microscopy. Collagen samples were treated with 0.02%, 0.04%, 0.10% and 0.20% glutaraldehyde individually at the first steps. Meanwhile, the other experimental groups were treated with 0.05%, 0.10%, 0.20% riboflavin, respectively, the structural change of the collagen network with 15, 30, 45, 60 min UV365 exposure and the fluorescent irradiation as the control group in turn was investigated by atomic force microscopy. The width and height of collagen assembly increased with the concentration of glutaraldehyde, which were changed from 36.45 ± 4.45nm to 64.35 ± 2.58nm and 2.26 ± 0.19nm to 4.93 ± 0.29 nm, respectively. The effect of different UV365 irradiation time on the height of collagen was greater than the effect on the width, the width and height of collagen were varied from 41.36 ± 4.30nm to 55.47 ± 4.53nm and 3.10 ± 0.39nm to 6.20 ± 0.85nm, which were changed 25.44% and 50.00%, respectively. The concentration of riboflavin was more efficiently than irradiation time, and the maximum width and height of collagen reached 106.98 ± 3.37nm and 11.17 ± 1.33 nm, respectively, the control group without a significant change at the fluorescent irradiation by the time. This optimized cross-linking method with riboflavin/UV365 would have great potential for tissue engineering in clinic and food packaging science.

Analysis of the Physico-Chemical, Mechanical and Biological Properties of Crosslinked Type-I Collagen from Horse Tendon: Towards the Development of Ideal Scaffolding Material for Urethral Regeneration

“…While collagen bands were clearly visible in the DHT film, no protein bands were detectable in the DHT/EDC film. The absence of collagen bands could be ascribed to the chemical EDC crosslinking that, by inducing zero-length crosslinks between the carboxylate moiety of aspartate and glutamate and amine residues of lysine, leads to the formation of high molecular weight aggregates that do not pass through polyacrylamide gel meshes [76,77]. Therefore, we assume that in our experiment, collagen subunits were highly crosslinked rather than degraded.…”

“…The electrophoretic pattern of collagen-based substrates confirmed that the applied crosslinking treatments did not induce collagen degradation. In particular, while α1(I) and α2(I) bands were clearly visible in the DHT film, no protein bands or smear were detectable in the DHT/EDC film because of the formation of high-molecular-weight crosslinked aggregates that did not pass through the polyacrylamide gel meshes [76,77,105]. This result was supported by AFM analysis, in which type I collagen native fibrillar structure with the typical D-banding periodicity was clearly visible for both DHT and DHT/EDC films.…”