Background: Porous silicon (pSi) nanoparticles (NPs) functionalized with suitable targeting ligands are now established cancer bioimaging agents and drug-delivery platforms. With growing interest in peptides as tumor-targeting ligands, much work has focused on the use of various peptides in combination with pSi NPs for cancer theranostics. Here, the authors investigated the targeting potential of pSi NPs functionalized with two types of peptide, a linear 10-mer peptide and its branched (Y-shaped) equivalent, that respond to legumain activity in tumor cells. Results: In vitro experiments established that the linear peptide-pSi NP conjugate had better aqueous stability under tumor conditions and higher binding efficiency (p < 0.001) toward legumain-expressing cells such as RAW 264.7 cells compared with that of its branched equivalent. In vivo studies (analyzed using ex vivo fluorescence) with the linear peptide-pSi NP formulation using a syngeneic mouse model of breast cancer showed a higher accumulation (p > 0.05) of linear peptide-conjugated pSi NPs in the tumor site within 4 h compared with nonconjugated pSi NPs. These results suggest that the linear peptide-pSi NP formulation is a nontoxic, stable and efficient fluorescence bioimaging agent and potential drug-delivery platform.