【摘要】：目的:建立HPLC-Protein A定量檢測(cè)貝伐珠單抗的方法,用于該單抗生產(chǎn)過(guò)程的含量測(cè)定和質(zhì)量控制。方法:采用Agilent Bio-Monolith Protein A Column(5.2 mm×4.95 mm,0.10 m L)色譜柱,以PBS(含0.12 mol·L-1氯化鈉,p H 7.4)為流動(dòng)相A,0.5 mol·L-1醋酸(p H 2.5)為流動(dòng)相B。上樣后用流動(dòng)相A沖平,然后用流動(dòng)相B進(jìn)行洗脫,流速為1.0 m L·min-1,柱溫為25℃,檢測(cè)波長(zhǎng)為280 nm。結(jié)果:以質(zhì)量濃度為橫坐標(biāo),峰面積為縱坐標(biāo)繪制標(biāo)準(zhǔn)曲線,在0.078~10.0 mg·m L-1濃度范圍內(nèi),R20.999;回收率在98.32%~101.7%之間;批內(nèi)精密度RSD≤1.5%;批間精密度RSD≤1.6%;檢測(cè)限0.30μg,定量限0.90μg;在p H變化±0.2、流動(dòng)相A中氯化鈉濃度變化±10.0%、柱溫變化±5.0℃、流速變化±20.0%條件下,峰面積相對(duì)標(biāo)準(zhǔn)差RSD為0.39%~1.0%;發(fā)酵液樣品的保留時(shí)間為1.350 min,與對(duì)照品的保留時(shí)間一致;在18 h內(nèi),同一批發(fā)酵液共檢測(cè)27次,峰面積的RSD為0.44%。結(jié)論:方法學(xué)驗(yàn)證結(jié)果顯示HPLC-Protein A法可以用于貝伐珠單抗生產(chǎn)過(guò)程的含量測(cè)定和質(zhì)量控制。
[Abstract]:Aim: to establish a method for the quantitative determination of bevacizumab by HPLC-Protein A, and to apply it to the determination and quality control of the mAb in the production process. Methods: a Agilent Bio-Monolith Protein A Column (5.2 mm 脳 4.95 mm,0.10 m L) column was used with PBS (containing 0.12 mol 路L-1 sodium chloride, p H 7.4) as the mobile phase A. 0.5 mol 路L-1 (p H 2.5) was used as mobile phase B. The sample was flushed with mobile phase A and eluted with mobile phase B, the flow rate was 1.0ml 路min-1, the column temperature was 25 鈩，

本文編號(hào)：2438538