Chronic pain is a major health problem worldwide but the limited knowledge of its underlying pathophysiology impairs the opportunities for diagnostics and treatment. Biomarkers of chronic pain are greatly needed to understand the disease and develop new targets for interventions and drug treatments, and potentially introduce more precise diagnostic procedures. Much evidence points to a neuroimmune pathology for many chronic pain conditions and that important neuroimmune biomarkers exist in the cerebrospinal fluid (CSF) of patients with chronic pain. Systematic collection of CSF in large cohorts of chronic pain patients and healthy volunteers has proven difficult, however. We established the Danish Pain Research Biobank (DANPAIN-Biobank) with the aim of studying potential neuroimmune and glia-related biomarkers of chronic pain. In this paper, we describe the methods and the study population of the DANPAIN-Biobank.