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Photoacoustic imaging is the latest promising diagnostic modality that has various advantages such as high spatial resolution, deep penetration depth, and use of non-ionizing radiation. It also employs a non-invasive imaging technique and optically functionalized imaging. The goal of this study was to develop a nanomedicine for simultaneous cancer therapy and diagnosis based on photoacoustic imaging. Human serum albumin nanoparticles loaded with melanin and paclitaxel (HMP-NPs) were developed using the desolvation technique. The photoacoustic-based diagnostic and chemotherapeutic properties of HMP-NPs were evaluated through in vitro and in vivo experiments. The size and zeta potential of the HMP-NPs were found to be 192.8 ± 21.11 nm and − 22.2 ± 4.39 mV, respectively. In in vitro experiments, HMP-NPs produced increased photoacoustic signal intensity because of the loaded melanin and decreased cellular viability because of the encapsulated paclitaxel, compared to the free human serum albumin nanoparticles (the control). In vivo experiments showed that the HMP-NPs efficiently accumulated inside the tumor, resulting in the enhanced photoacoustic signal intensity in the tumor site, compared to the normal tissues. The in vivo chemotherapy study demonstrated that HMP-NPs had the capability to treat cancer for an extended period. In conclusion, HMP-NPs were simultaneously capable of photoacoustic diagnostic and chemotherapy against cancer. [ABSTRACT FROM AUTHOR]