Photoacoustic detection is an emerging noninvasive and nonionizing detection technique with the merits of rich contrast, high resolution, and deep tissue penetration, especially for in vivo detection and imaging. Herein, we developed a photoacoustic (PA) molecular imaging probe (denoted as nanonaps) composed of a naphthalocyanine dye and a heptamethine dye as the internal standard with unchanged signals at 860 nm and the sensing component with peroxynitrite (ONOO-) target-decreased signals at 775 nm, respectively. The as-prepared nanonaps displayed high sensitivity and specificity of ONOO- both in vitro and in vivo. The PA860/PA775 ratio was increased as a function of the concentration of ONOO- (0-250 nM). More interestingly, our ratiometric nanonaps could be used for in vivo detection and imaging of ONOO-.