Selection of Protein Standards in Estimation of Total Protein Content in Commercially Available Intravenous Immunoglobulin (IVIG) Preparations by Biuret Method and its Comparison with Kjeldahl Method: Guidance to IVIG Manufacturers

Immunoglobulins (IgG) derived from the human plasma are used as bio therapeutics primarily in the form of polyclonal IgG or as hyper immune sera. Intravenous Immunoglobulin (IVIG) has been the driving force of the growth of plasma products industry. The manufacturing process of IVIG begins with collection & testing of single donation plasma, pooling of thousands of these donations, confirmation of plasma pool for freedom from viral markers, followed by very extensive process of separation and purification of the plasma pool with ethanol or chromatography. To assure the quality each batch of the product has to go through various quality control tests as mentioned in the Indian Pharmacopoeia monograph. The total protein content is directly linked to the efficacy of the treatment of patients having Primary Immune Deficiencies or other immunological disorders. The gold standard method of Kjeldahl for total nitrogen estimation has been specified in the pharmacopoeia but most of the time the manufacturers prefer to use biuret method for total protein estimation due to its ease of performance and comparative less time taken for obtaining the results. National Institute of Biologicals (NIB) being the notified Central Drug Laboratory for Blood products has been considered as the nodal organization in India to assess and verify test methods applicable for assuring the quality of blood products. The present study aims at assessing the suitability of biuret method without Trichloroacetic acid precipitation of proteins, for total protein estimation in the commercially available IVIG preparations and also to highlight various critical parameters to be considered to ensure reliable results which will be useful for the IVIG manufacturers for reliable assessment of the protein content of their product.