Liquid Transmission Electron Microscopy for Probing Collagen Biomineralization
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abstract
Collagen biomineralization is fundamental to hard tissue assembly. While studied extensively, collagen mineralization processes are not fully understood, with the majority of theories derived from electron microscopy (EM) under static, dehydrated, or frozen conditions, unlike the liquid phase environment where mineralization occurs. Herein, novel liquid transmission EM (TEM) strategies are presented, in which collagen mineralization was explored in liquid for the first time via TEM. Custom thin-film enclosures were employed to visualize the mineralization of reconstituted collagen fibrils in a calcium phosphate and polyaspartic acid solution to promote intrafibrillar mineralization. TEM highlighted that at early time points precursor mineral particles attached to collagen and progressed to crystalline mineral platelets aligned with fibrils at later time points. This aligns with observations from other techniques and validates the liquid TEM approach. This work provides a new liquid imaging approach for exploring collagen biomineralization, advancing toward understanding disease pathogenesis and remineralization strategies for hard tissues.
