A novel crosslinker-free technique toward the fabrication of collagen microspheres

Injectable collagen microspheres (CMs) have the potential to be an excellent tool to deliver various modulatory agents or to be used as a cellular transporter. A drawback has been the difficulty in producing reliable and spherical CMs. A crosslinker-free method to fabricate CMs was developed using liquid collagen (LC) in a water-in-oil emulsion process with varying concentrations of surfactant span-80. Different emulsion times of up to 16-hr were utilized to produce the CMs. Visual microscopy and scanning electron microscopy were utilized to determine the morphology of the CMs. To determine the fibril nature of the CMs, focus ion beam milling, energy dispersive spectroscopy, and Fourier Transformation-Infrared spectroscopy were performed. A cell biocompatibility study was performed to assess the biocompatibility of the CMs. The results demonstrated that consistent spherical CMs were achievable by changing the span-80 concentration. The CMs were fibrilized not only at the surface, but also at the core. Both the 1- and 16-hr emulsion time demonstrated biocompatibility and it appeared that the cells preferentially adhered to the CMs. This crosslinker-free method to fabricate CMs resulted in spherical, stable, biocompatible CMs, and could be an excellent technique for multiple tissue engineering applications.