Starch nanoparticles (sNPs) are considered ideal materials for applications in plant and agricultural sciences aiming at increasing crop yields, and improving resilience due to their non-toxicity, global availability, hydrophilicity, and biodegradability. However, the lack of research on the interaction between sNPs and plant cell walls has limited their application in these fields. Here, we designed Nile blue A-based sNPs (NB@G50-NPs) to investigate the penetration of small-sized sNPs (G50-NPs) through the plant cell wall. We demonstrate that 20 nm NB@G50-NPs can spontaneously cross the cell wall barrier and are rapidly taken up by Arabidopsis roots within a short time (30 min), showing nearly 10-fold higher fluorescence intensity compared to the free fluorescent dye. Additionally, the fluorescence quantum yield of NB@G50-NPs increases from 9% (for free dye) to 14%, and the particles show high stability across a broad pH range (3-10). This pH stability covers the entire pH range found in plant tissues. Our findings suggest that sNPs offer significant advantages for live cell imaging in plants and provide a foundation for future applications of sNPs in plant nanotechnology, including nanofertilizers, nanopesticides, and plant genetic engineering.