Braude I A
Prep Biochem. 1983;13(3):177-90. doi: 10.1080/00327488308064247.
A preparative, sequential chromatographic procedure has been developed for the purification of human gamma interferon (HuIFN-gamma). The four steps in the procedure are Controlled Pore Glass-adsorption chromatography, Concanavalin-A affinity chromatography, Heparin-Sepharose affinity chromatography and gel-filtration. By virtue of the development of a coordinated effluent-affluent buffer scheme, eluants can also serve as loading buffers for the succeeding column. Consequently, crude HuIFN-gamma preparations can be purified rapidly (approximately one week), easily, and is amenable to a semi-automated process. The procedure has also been shown to be efficient. Here, as an example, it is reported that an overall purification of greater than 75,000-fold can be achieved, yielding a specific activity of 5.2 X 10(7) units/mg, and a recovery of 95.5%. In addition, the peak fraction, representing 37.8% of the applied activity, had a specific activity of 1.0 X 10(8) units/mg protein and represents a purification of more than 145,000-fold. An SDS-PAGE analysis of one such fraction indicated that approximately 40% of the final material was HuIFN-gamma.
已开发出一种用于纯化人γ干扰素（HuIFN-γ）的制备性连续色谱方法。该方法的四个步骤为可控孔径玻璃吸附色谱、伴刀豆球蛋白A亲和色谱、肝素琼脂糖亲和色谱和凝胶过滤。由于开发了一种协调的流出液-流入液缓冲方案，洗脱液也可作为后续柱的上样缓冲液。因此，粗制的HuIFN-γ制剂能够快速（约一周）、轻松地得到纯化，并且适合半自动化操作。该方法也已证明是高效的。在此，作为一个例子，据报道可以实现超过75000倍的总纯化，比活性为5.2×10⁷单位/毫克，回收率为95.5%。此外，代表37.8%施加活性的峰馏分，其比活性为1.0×10⁸单位/毫克蛋白质，纯化倍数超过145000倍。对其中一个这样的馏分进行的SDS-PAGE分析表明，最终产物中约40%是HuIFN-γ。
