The development of recombinant DNA technology has provided a source of producing large quantities of biologically and medically important peptides and proteins. These peptides and proteins are produced in host organisms and isolated from cell extracts under conditions and in concentrations quite different than the molecule would ordinarily see under the physiological conditions in which it naturally occurs. Such conditions can in certain instances favor the formation of dimer, trimer and higher oligomers of the peptide or protein molecule through intermolecular disulphide linkages, other covalent bonds, or non-covalent interactions.
Since in many cases these dimers and higher oligomers have either lower or no biological activity or else have the potential of causing deterious-side effects, such as antibody induction in patients, if the peptide or protein composition were to be used therapeutically by parenteral administration it is important to have available a selective assay which can distinguish these higher molecular species when present in admixture with the monomeric form. In this way it would be possible to carry out quality control analysis of the peptide or protein composition to detect the presence of the oligomers in the first instance and if present, would serve as a simple monitoring assay to assist in the purification of the compositions from the oligomers.
Immunoassays, particularly radio- or enzymeimmunoassays are well known in the art and have proved to be very useful for the detection and assay of peptides and proteins. Recently, monocloncal antibodies have become available. These antibodies have the property of being able to recognize a single epitope of a peptide or protein.
A sandwich-type immunoassay employing monoclonal antibodies that recognize different epitopes of the target protein has recently been developed for human leukocyte interferon, Staehelin et al., Methods Enzymol. 79, 589 (1981). In such assay one antibody is bound to a solid support in a manner which allows the antibody to bind any protein which has the specific epitope recognized by the antibody. A second monoclonal antibody that is labeled with a detectable label such as .sup.125 I or peroxidase is used as a probe. The second monoclonal antibody has been raised against a second epitope on the protein and thus will bind to the protein which is already bound to the first monoclonal antibody.
Such a sandwich immunoassay cannot distinguish between the monomeric and the corresponding dimeric, trimeric and higher oligomeric forms of the protein because the second antibody will bind to each of these forms since they all will have the second epitope site available.