Sudan Black B Pretreatment to Suppress Autofluorescence in Silk Fibroin Scaffolds

Natural polymers are extensively utilized as scaffoldmaterialsin tissue engineering and 3D disease modeling due to their generalfeatures of cytocompatibility, biodegradability, and ability to mimicthe architecture and mechanical properties of the native tissue. Amajor limitation of many polymeric scaffolds is their autofluorescenceunder common imaging methods. This autofluorescence, a particularchallenge with silk fibroin materials, can interfere with the visualizationof fluorescently labeled cells and proteins grown on or in these scaffolds,limiting the assessment of outcomes. Here, Sudan Black B (SBB) wassuccessfully used prefixation prior to cell seeding, in various silkmatrices and 3D model systems to quench silk autofluorescence forlive cell imaging. SBB was also trialed postfixation in silk hydrogels.We validated that multiple silk scaffolds pretreated with SBB (hexafluoro-2-propanol-silkscaffolds, salt-leached sponges, gel-spun catheters, and sponge-gelcomposite scaffolds) cultured with fibroblasts, adipose tissue, neuralcells, and myoblasts demonstrated improved image resolution when comparedto the nonpretreated scaffolds, while also maintaining normal cellbehavior (attachment, growth, proliferation, differentiation). SBBpretreatment of silk scaffolds is an option for scaffold systems thatrequire autofluorescence suppression.

Automated summary

Natural polymers are widely used in tissue engineering and 3D disease modeling due to their cytocompatibility, biodegradability, and ability to mimic native tissue. However, autofluorescence of these polymeric scaffolds can hinder the visualization and assessment of fluorescently labeled cells and proteins. This study successfully utilized Sudan Black B (SBB) to eliminate autofluorescence in silk-based scaffolds, resulting in improved image resolution without affecting cell behavior.