Adhesions represent a major burden in clinical practice, particularly following abdominal, intrauterine, pericardial and tendon surgical procedures. Adhesions are initiated by a disruption in the epithelial or mesothelial layer of tissue, which leads to fibrin adhesion sites due to the downregulation of fibrinolytic activity and an increase in fibrin deposition. Hence, the metabolic events involved in tissue healing, coagulation, inflammation, fibrinolysis and angiogenesis play a pivotal role in adhesion formation. Understanding these events, their interactions and their influence on the development of post-surgical adhesion is crucial for the development of effective therapies to prevent them. Mechanical barriers, antiadhesive agents and combination thereof are customarily used in the battle against adhesions. Although these systems seem to be effective at reducing adhesions in clinical procedures, their prevention remains still elusive, imposing the need for new antiadhesive strategies.

Collagen type I is the most abundant extracellular matrix protein, and collagen type I supramolecular assemblies (e.g., tissue grafts, biomaterials and cell-assembled systems) are used extensively in tissue engineering and regenerative medicine. Many studies, for convenience or economic reasons, do not accurately determine collagen type I purity, concentration, solubility and extent of cross-linking in biological specimens, frequently resulting in erroneous conclusions. In this protocol, we describe solubility; normal, reduced and delayed (interrupted) SDS-PAGE; hydroxyproline; Sircol collagen and Pierce BCA protein; denaturation temperature; ninhydrin/trinitrobenzene sulfonic acid; and collagenase assays and assess them in a diverse range of biological samples (e.g., tissue samples; purified solutions or lyophilized materials; 3D scaffolds, such as sponges and hydrogels; and cell media and layers). Collectively, the described protocols provide a comprehensive, yet fast and readily implemented, toolbox for collagen type I characterization in any biological specimen.