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Gold nanoparticles (AuNPs) are increasingly used in applications across the biomedical domain, yet their long-term biodistribution and biocompatibility remain poorly understood. Conventional brightfield microscopy imaging techniques often fail to detect AuNPs due to optical diffraction limits and lack of chromogenic contrast. Understanding the biodistribution and ultimate fate of these nonbiodegradable NPs is crucial for further development of AuNP-based therapeutics and diagnostics. Here, we present a label-free multiphoton luminescence (MPL) imaging workflow that enables sensitive detection of AuNPs in liver histology sections, even 1 year after intravenous (IV) administration. MPL imaging exploits the intrinsic nonlinear optical properties of AuNPs to generate broadband emission under ultrafast pulsed laser excitation, enabling subcellular localization without exogenous labels while having the ability to rapidly image entire organ sections. The intrinsic, distinct broadband MPL emission produced by gold allows us to study these NPs in their biological context without extrinsic labels while also faithfully representing the surrounding tissue architecture via autofluorescence and second harmonic generation. We demonstrate that MPL imaging detects up to 98% more AuNP-positive regions than brightfield microscopy in challenging low-dose (1 nM) conditions and requires no modification of standard histology workflows. Correlative imaging with SEM–EDS confirms high spatial specificity (AUC = 0.955) of MPL for AuNP localization. Dose-dependent retention patterns were observed across liver tissue, and MPL analysis showed strong correlation with ICP–MS quantification. Importantly, histological and immunohistochemical analyses (Masson’s trichrome, CD3, and TUNEL) revealed no significant fibrosis, immune activation, or apoptosis in liver tissue at either low (1 nM) or high (10 nM) doses at 1 year post IV administration. These findings establish MPL imaging as a robust, label-free tool for long-term tracking of AuNPs in biological tissue and highlight its potential for improving biodistribution and safety assessments.