In the immunoblot, antigens coated on membranes are used as a solid phase in order to detect specific antibodies in patient samples. The test performance is either manual, semi- or fully automated. If a sample contains specific antibodies, these bind to the membrane-bound antigens. In the next step, an alkaline phosphatase (AP) labelled antibody (conjugate) is added, which binds to the specific antibodies. The alkaline phosphatase catalyses a colour reaction with the subsequently added nitro blue tetrazolium chloride/5-bromo-4-chloro-3-indolyl phosphate (NBT/BCIP). If specific antibodies are present in the patient sample, a dark line appears at the respective antigen position. The intensity of the resulting staining is proportional to the antibody concentration in the sample.