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Flexible and transparent substrates are emerging as low cost and easy-to-operate support for surface-enhanced Raman spectroscopy (SERS). In particular, in situ SERS detection approach for surface characterization in transmission modality can be efficiently employed for non-invasive analysis of non-planar surfaces. Here we propose a new methodology to fabricate a homogenous, transparent, and flexible SERS membrane by the assistance of a thin TiO 2 porous layer deposited on the PDMS surface, which supports the uniform loading of gold nanoparticles over large area. The substrate was first characterized for homogeneity, sensitivity and repeatability using a model molecule for SERS, i.e., 7-mercapto-4-methylcoumarin. Satisfactory intra-substrate uniformity and inter-substrates repeatability was achieved, showing an RSD of 10%, and an analytical sensitivity down to 10 nM was determined with an EF of 3.4 × 10 5 ± 0.4 × 10 5 . Furthermore, SERS detection of pyrimethanil (PMT), a commonly employed pesticide in crops for human consumption, was performed in situ, exploiting the optical transparency of the device, using both model surfaces and non-flat bio-samples. PMT contamination at the phytochemical concentration levels corresponding to commonly used infield doses was successfully detected on the surface of the yellow Ficus benjiamina leaves, supporting the use of this substrate for food safety in-field application.