Detection system unlabeled antibody

Multi-step technique (3) This is an indirect/direct method combining unlabeled primary antibodies with directly-conjugated antibodies. The method starts with staining the unlabeled antibody/antibodies with the appropriate detection system, but without performing the final enzymatic staining reaction. The tissue is blocked with normal serum from the host of the first primary antibody before the second, directly-labeled primary antibody is added. The staining ends with the two enzymatic reactions being performed sequentially. 

There are variety of sandwich ELISA systems. Eor example, in indirect sandwich ELISAs, the assay uses an unlabeled primary antibody for the sandwich step in conjunction with an enzyme-labeled anti-immvmoglobulin secondary antibody. This system can also employ a biotinylated sandwich antibody and enzyme-labeled streptavidin for detection. The indirect sandwich ELISA can be used not only for detection of antigens but also for measurement of antibody concentration, a system more specifically referred to as an indirect antibody capture immunoassay. In this case, an antigen-coated solid phase is prepared and then incubated with a serum or plasma test sample. Next, a secondary enzyme-labeled antiimmunoglobulin antibody or enzyme-labeled antigen is added, unbound reagents are removed by washing, and enzyme activity is assayed. 

---