T-2 toxin, one of the most dangerous natural pollutants, induces apoptosis through multiple pathways. Amongst, P53 mediated apoptosis pathway, an important collection of molecules, plays a key role in cell vital activity. Real-time monitoring of upstream and downstream activation relationships of P53 mRNA, Bax mRNA, and cytochrome c (Cyt c) in signaling pathways is of great significance for understanding the apoptotic machinery in human physiology. In this work, a novel nucleic acid multicolor fluorescent probe, based on silica-coated symmetric gold nanostars (S-AuNSs@SiO₂), was developed for highly sensitive in situ real-time imaging of P53 mRNA, Bax mRNA, and Cyt c during T-2 toxin-induced apoptosis. The nucleic acid chains modified with carboxyl groups were modified on the surface of S-AuNSs@SiO₂ by amide reaction. The complementary chains of targeted mRNA and the aptamer of targeted Cyt c were modified with different fluorophores, respectively, and successfully hybridized on S-AuNSs@SiO₂ surface. When targets were present, the fluorescent chains bound to the targets and detached from the material, resulting in the quenched fluorescence being revived. The probes based on S-AuNSs showed excellent performance is partly ascribed to the presence of 20 symmetric “hot spots”. Notably, the amide-bonded probe exhibited excellent anti-interference capability against biological agents (nucleases and biothiols). During the real-time fluorescence imaging of T-2 toxin-induced apoptosis, the corresponding fluorescence signals of P53 mRNA, Bax mRNA, and Cyt c were observed sequentially. Therefore, S-AuNSs@SiO₂ probe not only provides a novel tool for real-time monitoring of apoptosis pathways cascade but also has considerable potential in disease diagnosis and pharmaceutical medical.