Biopharmaceuticals (or biologics) are a class of drugs synthesized from biological sources that have a therapeutic effect. Monoclonal antibodies (mAbs) have become as one of the major biopharmaceuticals due to their high specificity to target molecules, thus reducing the risk of side effects to the patients. However, the cost of production of monoclonal antibodies is significantly higher than that of small molecule drug manufacture due to stringent purity regulatory requirements. Affinity column is normally used (such as protein A) to selectively bind antibodies in complex solutions allowing impurities to flow through. When the dynamic capacity of the column is reached and antibody begins flowing through the column without binding creates the breakthrough curve. Monitoring the breakthrough is very important to prevent precious antibody going to waste and also to save time by letting the operator know when to switch columns in a multi column format. Antibody breakthrough cannot be measured using UV absorbance because host cell proteins in the flow-through impede UV detection of antibody breakthrough. Other methods have been used such as enzyme-linked immunosorbent assay (ELISA), surface plasmon resonance (SPR), proximity based homogeneous assay, Raman spectroscopy, etc. However, these technologies are generally too slow and/or require too many reagents. The present invention disclosure uses fluorescence polarization/anisotropy based for determining low level monoclonal antibody from the effluent of chromatography column.