The effect of cross-linking time on a porous freeze-dried collagen scaffold using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide as a cross-linker.

Collagen is a widely studied natural polymer for use as a scaffold material for various tissue engineering applications. Untreated collagen, however, has a fast biodegradation rate and low mechanical strength. Additionally, collagen cross-linking has been studied extensively and different cross-linking agents and methods have been explored. Although glutaraldehyde (GA) is the most convenient and traditional chemical agent currently used for this purpose, it nevertheless poses potential cytotoxicity. Therefore, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) is widely being studied as well, due to its characteristic of being a zero-length cross-linker that does not remain in the collagen's structure. Nevertheless, cross-linking with EDC can be implemented in several ways. In this study, two methods of cross-linking with EDC, prior to and post freeze-drying, were examined for freezedried collagen scaffolds. Four different collagen concentrations between 0.3 and 2.0 w% were inspected and different cross-linking methods were examined by a differential scanning calorimeter (DSC) to determine the collagen's denaturation temperature (Td). Furthermore, the water uptake abilities of the scaffolds were tested in phosphate buffered saline (PBS) and the scaffolds' pore structure was examined with a scanning electron microscope (SEM). As assumed, the DSC measurements demonstrated that collagen Td values increased with increasing collagen concentration. With lower collagen concentrations, the cross-linking post freeze-drying enhanced the Td values, but with higher collagen concentrations, the Td values were increased only with cross-linking prior to freeze-drying. The water uptake measurement showed increased water uptake ability when cross-linking post freezedrying. The pore sizes of the different collagen scaffolds were between 50 and 120 mum.