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The objective of this paper is to evaluate the penetration and accumulation of SiO₂ nanoparticles (NPs) in vitro human placental tissue as a function of NP size and electroporation (EP) and microneedle (MN) treatments. The results show that the penetration and accumulation of SiO₂ NPs has a significant influence on the optical properties. Optical coherence tomography (OCT) monitoring and diffuse reflectance (DR) spectra measurements indicate that 30 nm SiO₂ NPs have a tendency to generate greater signal intensities, while 15 nm NPs penetrated faster and had a higher penetration depth. The reduction in average attenuation coefficients of human placental tissue when the SiO₂ NPs size is constant is found to show the following trend: SiO₂ NPs in combination with EP and MN treatment > SiO₂ NPs in combination with MN > SiO₂ NPs with EP > only SiO₂ NPs. It can be concluded that EP and MN treatments dramatically increase the penetration of SiO₂ NPs into human placental tissue. The results prove that OCT and DR spectra can be combined with EP and MN treatments to establish a theoretical basis for the safe use of SiO₂ NPs on human placental tissue and can potentially become a powerful tool for early diagnosis and monitoring of tissue diseases.