Astrocytes are intimately involved in immunological and inflammatory events occurring in the central nervous system (CNS), due to their ability to secrete and respond to a large number of immunoregulatory cytokines/chemokines such as IL-1β, IL-6, IL-8, IL-10, IL-17, IL-27, TNF-α, TGF-β, IFN-γ, IFN-β, CCL2, CCL3, CCL5, CXCL10, and CXCL12. Although expression of cytokines and chemokines is limited in the normal CNS, elevated expression of these proteins, as seen in disease entities such as multiple sclerosis (MS), HIV-1 associated neurocognitive disorders (HAND), Alzheimer’s disease (AD), Parkinson’s disease (PD) and amyotrophic lateral sclerosis (ALS), contributes to the development of inflammation and neuronal demise in these diseases. As a potent source of cytokines and chemokines, astrocytes play a pivotal role in the type and extent of neuroinflammatory responses. Astrocytes can be stimulated in vitro to produce numerous cytokines/chemokines, which are secreted and detected in supernatants by a technique known as enzyme-linked immunosorbent assay (ELISA). In this chapter, we describe our experience using ELISAs to detect and quantify cytokines and chemokines secreted by stimulated murine astrocytes, specifically IL-6 and CXCL10.