Visualizing Endogenous Amyloids in Spermatozoa

Endogenous amyloids from fractionated or unfractionated SP were visualized by staining samples with Proteostat and/or pFTAA, using approaches previously described (Usmani et al., 2014 (link)). Briefly, fractionated or unfractionated SP were incubated with Proteostat and/or pFTAA for 15 min at room temperature, and then transferred into an Ibidi Chamber Slide (#80826 from GmbH). Stained samples were imaged on a Zeiss LSM710 AxioObserver confocal microscope equipped with a Plan-Apochromat 63/1.40 oil objective lens and Zen-Software v2010 (Zeiss, Germany). Spermatozoa were simultaneously imaged by DIC using transmitted light detector (T-PMT) and appropriate condenser settings. Where indicated, spermatozoa were additionally visualized by staining with Hoechst 33342.