The understanding of disease mechanisms and the search for potential therapeutic targets have driven the focus of proteomics studies towards the identification of specific protein profiles and molecular pathways disease associated. Protein microarrays offer high-throughput alternatives to mass spectrometry, with peptide, phage, and recombinant protein microarrays providing different options for studying disease-specific alterations. Commercially available antibody microarrays, particularly cytokine arrays, enable the simultaneous analysis of multiple cytokine expression levels, offering valuable insights into disease processes, drug efficacy, and biomarker discovery. Indeed, cytokine arrays outperform mass spectrometry methods due to their ability to analyze cytokines at an extremely low concentration in parallel in a high-throughput manner. Here, we outline protocols for cytokine analysis using cytokine antibody microarrays, detailing sandwich approaches and direct or indirect incubation methods for the characterization of disease-associated cytokine profiles.