Porous silicon nanoneedles can map Cathepsin B activity across normal and tumor human esophageal mucosa. Assembling a peptide-based Cathepsin B cleavable sensor over a large array of nano­needles allows the discrimination of cancer cells from healthy ones in mixed culture. The same sensor applied to tissue can map Cathepsin B activity with high resolution across the tumor margin area of esophageal adenocarcinoma.