Introduction
The quality of Ligand Binding Assays (LBAs) is dependent on the use of quality Critical Reagents (CR). Typically, conjugations are performed by a procedure of buffer exchange, conjugation and a final buffer exchange using molecular weight cut-off or desalting spin columns. Incorporation is then determined by a wet chemistry method (e.g. HABA). Many of these procedures are not well controlled and do not ensure lot-to-lot consistency. By utilizing preparative SEC, High Resolution Mass Spectrometry (HRMS), analytical SEC (aSEC), and Tangential Flow Filtration (TFF), we have developed a method for ensuring uniformity of incorporation ratio, purity and concentration throughout the life cycle of a CR. This effectively eliminates the need for tedious and costly assay bridging and perhaps, revalidation.
