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    • 3. 发明授权
    • Purified ciliary neurotrophic factor
    • 纯化的睫状神经营养因子
    • US5011914A
    • 1991-04-30
    • US293851
    • 1989-01-05
    • Franklin D. CollinsLeu-Fen Lin
    • Franklin D. CollinsLeu-Fen Lin
    • A61K38/00C07K14/475C07K16/22
    • C07K14/475C07K16/22A61K38/00
    • A ciliary neurotrophic factor (CNTF), particularly sciatic nerve CNTF (SN-CNFT) is claimed. The SN-CNTF described herein is a single protein species and has a specific activity that increased to greater than 25,000-fold from crude extract. The purification is carried out by lowering the pH of the crude nerve extract preparation to form a precipitate which is removed and discarded; raising the pH to about 6.3 followed by ammonium sulfate fractionation; chromatofocusing a solution containing a second precipitate obtained from the 30% to 60% ammonium sulfate containing solution; subjecting the fractions obtained by chromatofocusing to sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE); and performing reversed-phase high-performance liquid chromatography (HPLC) on the SDS-PAGE eluate.Amino acid data for this SN-CNTF is also provided. In addition, methods for using this data for providing SN-CNTF probes and for screening cDNA and genomic libraries are also provided. Recombinant-DNA methods for the production of SN-CNTF are described.
    • 要求提供睫状神经营养因子(CNTF),特别是坐骨神经CNTF(SN-CNFT)。 本文所述的SN-CNTF是单一蛋白质物质,并且具有比粗提取物增加至大于25,000倍的比活性。 通过降低粗神经提取物制剂的pH以形成除去并丢弃的沉淀物来进行纯化; 将pH升高至约6.3,随后进行硫酸铵分级; 将含有30%至60%含硫酸铵的溶液得到的含有第二沉淀的溶液进行色谱聚焦; 将通过色谱聚焦获得的级分进行十二烷基硫酸钠(SDS) - 聚丙烯酰胺凝胶电泳(PAGE); 并在SDS-PAGE洗脱液上进行反相高效液相色谱(HPLC)。 还提供了该SN-CNTF的氨基酸数据。 此外,还提供了使用该数据提供SN-CNTF探针和筛选cDNA和基因组文库的方法。 描述了用于生产SN-CNTF的重组DNA方法。