The tumour microenvironment (TME) is a complex system that contains both tumour cells and lymphocytes that have infiltrated a patient's tumour. These lymphocytes can have both antitumorigenic and pro-tumorigenic properties within the complex nature of the TME, also often contributing to tumorigenesis. The lymphocytes within the TME are often made up of a significant number of bystander cells, cells that have little or no mmunogenic effect (Meier and Satpathy, 2022). Single cell analysis and T cell therapy have revolutionised the field of cancer treatment and immunotherapy by providing a personalised approach to patient care. Single cell analysis allows researchers to gain a deeper understanding of the heterogeneity within a tumour and identify specific cells within the tumour microenvironment that may be a potential therapeutic. In this study the development and optimisation of a process that uses RemedyBio’s (the company) Nanoreactor single cell analysis platform and functionalised PDMS assays to screen a patient resected tumour for those rare functionally relevant TILs is discussed. The overall hypothesis is that identifying and expanding only the small subpopulation of TILs that are immunologically functional against a patient’s cancer could lead to a more robust and potent therapeutic. Almost 2 million single T cells were screened using fluorescent based PDMS assays that were developed using both healthy donor T cells and patient TILs from dissociated tumour samples. The process was then further verified using a number of different known target screening methods to ensure the analytical tool, was capable of screening patient samples and detecting cytokines such as IFN-γ and granzyme B at low concentrations from a single cell. Gold standard methods such as ELISA and FACS were also used to assess the TIL population being screened. Analysis of millions of cells from patient TME simultaneously at a multiplexed level offers the potential to develop a more potent, yet less harmful therapeutic for those with currently incurable cancers. Prior to commencing this research, it was unknown if functional cytokine markers for T cell activation and cell killing could be detected from the secretome of a single cell. Initial data shows that IFN-γ and granzyme B can be detected in low concentrations from single cells within a microcapillary array using a sandwich immunoassay with a fluorescent based detection method on APTES treated PDMS.