Identifying cytotoxic T-cells in cleared, thick tissue samples through CD8 Staining

Human tonsil stained with Biocare Medical SP6 in a 1:100 concentration.

Formalin-fixed, paraffin-embedded human tonsil tissue ranging in size up to 3mm was de-waxed, pretreated, and subject to antigen retrieval using an EDTA/Methanol system.  The tissue was then stained using Biocare Medical anti-CD8 (SP6) and a nuclear stain, TO-PRO-3 (Thermo-Fisher). The tissue was cleared using a hybrid iDISCO protocol before imaging on Alpenglow Biosciences Aurora™ 3Di Hybrid Open Top Light Sheet (HOTLS) fluorescent microscope. For more information and a list of optimized antibodies for deep tissue imaging, see our brochure “Staining with Immunomarkers within Deep Tissues”.