An improved anion-exchange HPLC method for the detection and purification of adenoviral particles

We have developed an anion-exchange high-performance liquid chromatography (HPLC) method using Q Sepharose XL (Amersham Pharmacia Biotech) as adsorben t to analyze samples containing adenovirus. This method has several major a dvantages over the HPLC method previously described for quantitating partic les, namely (1) a > 10-fold improvement in the detection limit of adenoviru s in crude preparations; (2) absence of interferences originating from nucl eic acids and proteins which usually contaminate crude samples; (3) unprece dented sharpness and symmetry of adenovirus peak, rendering the identificat ion of the viral peak unambiguous, even in extremely crude and dilute prepa rations; and (4) no enzymatic treatment required even for crude samples. Th is assay was used to quantitate particles in samples taken at the transfect ion and amplification stages of production of various recombinant adenoviru s, and in cultures of wild-type adenovirus of different serotypes. A modifi cation of this analytical method was also developed for the purification of infectious adenovirus particles, including fiber-modified and third-genera tion recombinant viruses, giving highly purified preparations from low-tite r crude lysates with an excellent overall recovery (50-74%).