Determination of percentage antigen adsorption in vaccine drug formulation.

疫苗药物配方的生产是一项价值数百万美元的商业活动。要实现工业化生产，通常需要制造高效率的药物产品，这通常是通过在药物配方中添加佐剂来获得的，但不同的药物配方对抗原的吸附百分比差别很大。由于免疫原性与抗原吸附百分率直接相关，因此开发一种定量测定最终药物产品中吸附抗原的方法是非常关键的。抗原吸附百分比的测定需要对药物产品中的游离抗原和吸附抗原进行量化。赛诺菲巴斯德正在尝试开发一种使用水相结合LC-MS法定量测定游离抗原的方法。目前正在对该方法进行优化，以克服基质效应。吸附抗原的定量仍然是一个挑战，我们建议使用不同的试剂和温度条件解吸抗原，以破坏抗原和佐剂之间的强烈静电相互作用。药物产品将被离心，以分离游离抗原。将含有吸附抗原的小球重新悬浮，并用十六烷基吡啶氯化物、柠檬酸二水合钠、磷酸二钠、盐酸胍、尿素和琥珀酸试剂处理。一旦抗原被成功解吸，游离的抗原将被胰酶消化，并用赛诺菲巴斯德开发的LC-MS方法进行定量。这种方法将是制药研究的一项进步，将提高患者的药物疗效，并提高加拿大的研究在世界科学界的知名度和认知度。

Production of vaccine drug formulation is a multi-million dollar commercial activity. To operate on an industrial scale normally requires the manufacturing of high efficiency drug products which is typically obtained by the addition of adjuvants in drug formulations, but percentage adsorption of antigens vary dramatically among different drug formulations. Since immunogenicity is directly linked with percentage antigen adsorption, it is extremely critical to develop a method for the quantification of adsorbed antigens in the final drug product. Determination of percentage antigen adsorption would require the quantification of both free and adsorbed antigens in the drug product. Sanofi Pasteur is attempting to develop a method for quantification of free antigens using AQUA method coupled with LC-MS. The method is currently being optimized to overcome the matrix effect. The quantification of adsorbed antigens is still a challenge and we propose to desorb antigens using various reagents and temperature conditions to breakdown the strong electrostatic interactions between antigens and adjuvant. The drug product will be centrifuged to isolate free antigen. The pellet containing adsorbed antigen will be resuspended and treated with reagents such as cetylpyridinium chloride, sodium citrate dihydrate, sodium phosphate dibasic, guanidine HCl, urea and succinate. Once the antigens are successfully desorbed, the free antigens will be digested with trypsin and quantified by LC-MS method being developed by Sanofi Pasteur. This method will be an advancement in pharmaceutical research that will enhance drug efficacy in patients and bring increased visibility and recognition to Canadian research within the world-wide scientific community.