Collagen fibrils were in vitro reconstituted from the aggregated collagen solution in the presence of a wide range of collagen/chondroitin 4-sulfate (Col/C4S) ratios. As revealed by turbidimetric measurement, the collagen fibril formation is significantly accelerated by C4S. The turbidity values of both the solution after 30 min preincubation at 4°C and the gels after 60 min preincubation at 37°C become larger with the increase of C4S amount. According to the results obtained from turbidimetric measurement and atomic force microscopy observation of solutions, it is predicted that the preincubation of Col/C4S blends at 4°C is necessary to initiate the Col/C4S binding and then promote the further lateral fusion of collagen aggregates in solution. The interactions between collagen and C4S are also vital in the growth phase of collagen self-assembly. Collagen quantitation data show that the amounts of collagen incorporated into self-assembled cofibrils increase a lot as a result of the presence of C4S. Differential scanning calorimetry measurement shows that the thermal stability of cofibrils keeps increasing with the ascending amount of incorporated C4S. It is suggested that the bound C4S might be captured inside the cofibrils acting as promoter and stabilizer. Atomic force microscopy and scanning electron microscopy observations of self-assembled fibrils indicate that the size increase of the self-assembled cofibrils depends on the lateral accretion of small collagen fibrils, while the self-assembly mode of collagen is not affected.
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