Neocuproine is widely used as a chelating agent in analytical chemistry, particularly for the determination of copper ions. It forms a stable complex with copper(I), which is utilized in spectrophotometric assays to measure copper concentrations in various samples, including environmental and biological specimens.

In biochemistry, neocuproine is employed to study copper-dependent enzymes and proteins, helping to understand their mechanisms and interactions. It is also used in antioxidant assays to quantify the reducing capacity of substances by measuring the reduction of copper(II) to copper(I).

Additionally, neocuproine finds application in organic synthesis as a catalyst or ligand in copper-mediated reactions, enhancing the efficiency and selectivity of certain chemical transformations. Its ability to stabilize copper in the +1 oxidation state makes it valuable in various research and industrial processes.