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In this study, we investigate the efficacy of optical photothermal infrared (O-PTIR) spectroscopy, also known as mid-infrared photothermal (MIP) microscopy, for label-free and nondestructive detection of micro- and nanoplastics (MNPs) down to diameters of 200 nm in mammalian tissues. Experiments with both in vitro three-dimensional cell cultures derived from HTC116 colorectal cancer cell line and in vivo mouse tissue models were conducted. Spherical polystyrene particles served as reliable model systems for evaluating spatial resolution limits and quality of spectra. Our findings demonstrate the superior resolution of O-PTIR in imaging individual particles of 200 nm in mouse kidney tissues, surpassing the capabilities of traditional Fourier transform infrared (FTIR) spectroscopy. Furthermore, we apply a semiautomated image analysis that incorporates machine learning algorithms to accelerate the detection process, thus improving throughput and minimizing the potential for human error. The results confirm that O-PTIR is able to provide high-quality, artifact-free spectral images in a contact-less manner and significantly outperforms traditional infrared spectroscopy in terms of spatial resolution and signal-to-noise ratio in complex biological matrices.