Crosslinking amine-rich biomolecules with aldehydes, such as paraformaldehyde (PFA) and glutaraldehyde (GA), is a widely used method for biomaterial fabrication and cell fixation. In this study, we introduce homobifunctional photocaged 2-butene-1,4-dials (bis-PBDAs) as novel amine-reactive photo-crosslinkers. Compared to PFA and GA, bis-PBDAs are inherently less toxic. Upon photo-deprotection, bis-PBDAs efficiently react with the amino groups of biomolecules, such as carboxymethyl chitosan and histone H3, forming stable, well-defined crosslinks. Even at low concentrations (~100 μM), bis-PBDAs facilitate effective cell fixation without significantly altering cellular morphology or interfering with biochemical analyses, including protein/mRNA extraction and immunofluorescence imaging. Therefore, bis-PBDAs are promising reagents for crosslinking amine-rich biopolymers and hold great potential in biomaterial fabrication and the fixation of living cells and tissues.
Keywords: Amine-rich biomolecules; Cell fixation; Hydrogel; Photo-crosslinker.
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