Process-specific assays utilize those host cell antigens most likely to copurify with the protein of interest, whereas generic assays use all recoverable host cell antigens, thereby testing for the presence of any antigen during protein production and that might therefore be recovered with the purified protein. Various assays to quantify these antigens in both in-process streams and purified bulk solutions have been described (1–3). The production of recombinant proteins for therapeutic and vaccine uses necessitates the analysis of purified products for residual host cell antigens. Other groups have reported on the utility of gel or fluorograph densitometry, using multiple detection methods, in carbohydrate analysis (5), taxonomic and forensic applications (6), as a clinical diagnostic tool in Balkan nephropathy (7) and in the analysis of high density lipoproteins (8), as a means of quantifying relative levels of excretory-secretory polypeptides synthesized in vitro by Schistosoma mansoni daughter sporocysts (9), and for analyzing adsorption of proteins and an oligodeoxynucle-otide in Alhydrogel®-based malaria vaccine candidates (10). This method was shown to be at least 20 times more sensitive than a commercially available ELISA kit designed to measure host cell impurity levels (3). reported a quantitative densitometric method that measures host cell derived impurity levels on immunoblots of recombinant proteins expressed in E. In addition, Morçöl and Subramanian reported the development of a sensitive Ponseau-S-stained dot blot/densitometric protein assay that was less subject to interference by pH, detergent and other reagents than the commonly used Bradford protein assay (2). For example, Vuletich and Osawa showed that densitometry following SDS-PAGE and electroblotting was 20-fold more sensitive than HPLC at detecting oxidatively modified myoglobin (1). In addition to being accurate, sensitive and reproducible, the technique is cost-effective, simple, and does not require a high degree of specialized training, yet provides technical advantages over other available tools. Densitometry is particularly useful due to its sensitivity, accuracy and versatility, and it can be applied to proteins in gels or on membranes and used with numerous detection methods. A variety of techniques are available for the quantification of proteins, their degradation products and other impurities.
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