Extraction and Characterization of Collagen from the Skin of <I>Arothron</I> <I>stellatus</I> Fish—A Novel Source of Collagen for Tissue Engineering

Synthesis of highly interconnected 3D scaffold from Arothron stellatus skin collagen for tissue engineering application

“…Amide I (1600-1660 cm -1 ) band associated with the carboxyl group within the triple helix, corresponding to 1651 cm -1 , plays an essential role in the secondary structure of the protein. The amide II (~1550 cm -1 ) at 1530 cm -1 and amide III (1320-1220 cm -1 ) at 1253 cm -1 mainly correspond to the N-H bending and N-H stretching vibration, respectively [7]. The triple helix structure of the collagen can be confirmed with respect absorption intensity to amide III and II, which was approximately equal to 1.0, in COL-SPG scaffold as reported by Gordon et al, 1974 andSylvestert et al, 1989 [11-13].…”

“…Type I collagen was extracted from AsFS and purified by the procedure reported earlier [7]. Dulbecco's Modified Eagle's Medium (DMEM), fetal bovine serum (FBS), and supplementary antibiotics for tissue culture were purchased from Sigma-Aldrich, Bangalore, India.…”

“…The surface morphology of the 3D COL-SPG was visualized using SEM (VEGA3 SBH TESCAN) operating at an accelerating voltage of 5-20 kV [7].…”

“…The NIH 3T3 fibroblast cell line and human keratinocytes cell line (HaCaT) were grown on the COL-SPG scaffold placed in 24-well plates (Corning, NY) and maintained in DMEM with 10% fetal calf serum supplemented with penicillin (120 units per mL), streptomycin (75 mg/mL -1 ), gentamycin (160 mg/mL -1 ), and amphotericin B (3 mg/mL -1 ) at 37°C at a density of 5 × 10 4 cells/mL and then incubated over a time interval for 24 h, 72h and 7 th day in a humidified atmosphere of 5% CO 2 . Cells cultured in blank wells were used as control [7]. After 24h, 72h and 7 th day the culture medium was replaced with a serum-free medium containing 10 µL of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and incubated at 37°C for 4 h in a humidified atmosphere of 5 % CO 2.…”

“…In TGA, it shows that the first weight loss below 100°C was due to the traces of moisture and the second weight loss at 330°C was due to the thermal destruction of the residual group and polypeptide chain in the collagen. Differential thermogravimetric analysis (DTA) clearly corresponds to the derivative weight percentage of the COL-SPG sample (Figure 1(c) to (d)) [7].…”