A critical driver in the development of single-cell analysis platforms has been the recognition that cellular heterogeneity is crucial to understanding disease. Single cell proteomics offer significant insights of cellular function; however, currently suffers from low-throughput. The work outlined here presents the development and application of several single-cell protein analysis systems. Each aim to address technological gaps regarding throughput, cell selectivity and amenability to processing samples directly from patients. To achieve higher-throughput, we have developed the CellWell platform, a high-density microwell array which can capture thousands of cells within minutes; however, posed challenges relating to the simultaneous lysis of these cells. We developed a facile method to produce surface microelectrodes to achieve single-cell lysis on-chip, but the demanding surface chemistry requirements imposed by the necessity to support simultaneously both the microelectrodes and single-molecule antibody microarrays proved difficult to overcome. Instead, we investigated how implementing semi-permeable hydrogel-based microwells could overcome these issues. To assay cells in patient blood samples with the CellWell, pre-processing is necessary. With a clinical setting in mind, it would be advantageous to process raw samples directly from patients with little or no off-chip pre-processing. To address this, we develop our methodology into the Hydrodynamic Trapping Centrifugal Release (HTCR) chip which is specifically designed to isolate cancer cells from patient liquid biopsies. The HTCR implements a method by which cells can be easily released from hydrodynamic traps and subsequently moved to isolated compartments. We conclude in validating the single-molecule single-cell method using fluorescence and immunofluorescence microscopy. While necessary to validate our single-molecule approach, we also show that the equivalence of these measurements of the steady-state distribution of protein abundance can be exploited to pave the way for absolute quantitation by fluorescence and immunofluorescence microscopy.