Abstract Introduction and Objectives: Despite multiple resections and long-term chemo and immunotherapy, most non-muscle invasive bladder cancer patients suffer from recurrence or progression leading to cystectomy and a less favorable outcome. Possible reasons for that are incomplete resection and reimplantation of cancer cells, which could be prevented by improved resection and adjuvant therapy. Our objective was to develop a targeted drug for detection, fluorescence-guided resection, and deep-penetrating adjuvant photodynamic therapy of urothelial carcinoma (UC). The agent was based on upconversion nanoparticles (UCNP), which can carry a photosensitizer and can transform deep-penetrating near-infrared light into high-energy visible light, demanded for tumor visualization and for production of reactive oxygen species in the photosensitizer. At this stage, we aimed to select an antibody that could be attached to UCNPs to deliver them to UC cells. Methods: We produced silica-coated UCNP of the composition NaYF4:Yb,Er amenable for conjugation with biomolecules. An anti-Glypican-1 (GPC-1) monoclonal antibody MIL-38 (Glytherix Ltd., Sydney, Australia), was chosen for targeted delivery of the nanoparticles as it had previously demonstrated affinity towards bladder cancer. UCNPs were conjugated with MIL-38 by using a fusion protein Linker-Protein G (LPG). Finally, to investigate targeted binding and molecular specificity of these nanoconjugates, we incubated them with GPC-1 positive and GPC-1 negative cells. The role of MIL-38 in targeted delivery of nanoconjugates was also validated by incubation of GPC-1 positive T24 cells with nanoparticles coupled to an isotype control antibody and without an antibody. Results: Targeted upconversion nanoconjugates UCNP-LPG-MIL-38 labeled almost 90% of T24 cells with high expression of GPC-1 and only 23.2% of C3 cells with low expression of this antigen, demonstrating high molecular selectivity and specificity. Incubation of T24 cells with nanoconjugates linked with a control antibody and without antibody resulted in labeling of 19.8% and 26.2%, respectively, demonstrating the role of MIL-38 in targeted delivery of these nanoconjugates. As a result of the labeling, mean photoluminescence of cells in targeted group was from five to eight times stronger than in control groups, allowing for easy identification of positive cells with low background autofluorescence. Conclusions: These results highlight the potential of these nanoconjugates for the diagnosis and therapy of UC, as they can bind to Glypican-1-positive BC cells and cause their bright photoluminescence, which could be used for detection of tumors and activation of photosensitizers. It was also confirmed that monoclonal antibody MIL-38 has high potential to be applied in experimental diagnosis, drug delivery, and targeted therapy of UC, as it mediated targeted binding of upconversion photoluminescent nanoconjugates to Glypican-1-positive UC cells. Citation Format: Liuen Liang, Andrew Care, Anwar Sunna, Douglas Campbell, Bradley Walsh, Andrei Zvyagin, David Gillatt, Dmitry Polikarpov. Photoluminescent nanoconjugates for molecular imaging of bladder cancer [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2019 May 18-21; Denver, CO. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(15_Suppl):Abstract nr B27.