Simultaneous High-Throughput Conformational and Colloidal Stability Screening Using a Fluorescent Molecular Rotor Dye, 4-(4-(Dimethylamino)styryl)-N-Methylpyridinium Iodide (DASPMI)
Simultaneous High-Throughput Conformational and Colloidal Stability Screening Using a Fluorescent Molecular Rotor Dye, 4-(4-(Dimethylamino)styryl)-N-Methylpyridinium Iodide (DASPMI)
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DOI：
10.1177/1087057116646553
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发表时间：
2016-09-01
影响因子：
--
通讯作者：
Katayama, Derrick S.
中科院分区：
文献类型：
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作者：
Wong, Jensen J. H.;Wright, Sara K.;Katayama, Derrick S.
用于提高筛选蛋白质制剂的通量的技术正在不断发展。本文的目的是突出的分子转子染料，4-（4-（二甲氨基）苯乙烯基）-N-甲基吡啶碘（DASPMI）在筛选蛋白质结构的构象稳定性，胶体稳定性和微妙的预转变动力学在早期配方开发的新应用。测量了单克隆抗体在吐温80存在下的表观解折叠温度（T-m），并将数据与差示扫描量热法（DSC）测量结果进行比较。此外，测量DASPMI的荧光强度作为蛋白质浓度的函数显示与通过DLS测量的两种不同单克隆抗体制剂的扩散相互作用参数（k（D））的一致相关性。最后，由于分子转子染料对微粘度（（微））变化的敏感性，两种单克隆抗体制剂与红边激发位移（REES）实验结果相关的微妙的预转变动力学是可辨别的。分子转子染料的这种新应用提供了一种有价值和有前途的方法，用于简化早期制剂开发过程，因为在96孔板格式中材料消耗低，分析时间快。
Technologies to improve the throughput for screening protein formulations are continuously evolving. The purpose of this article is to highlight novel applications of a molecular rotor dye, 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide (DASPMI) in screening for the conformational stability, colloidal stability, and subtle pretransition dynamics of protein structures during early formulation development. The measurement of the apparent unfolding temperature (T-m) for a monoclonal antibody in the presence of Tween 80 was conducted and data were compared to the results of differential scanning calorimetry (DSC) measurements. Additionally, measuring the fluorescence intensity of DASPMI as a function of protein concentration shows consistent correlation to the diffusion interaction parameter (k(D)) for two distinct monoclonal antibody formulations measured by DLS. Lastly, due to the sensitivity of the molecular rotor dye to changes in microviscosity ((micro)), subtle pretransition dynamics were discernable for two monoclonal antibody formulations that correlate with findings by red-edge excitation shift (REES) experiments. This novel application of molecular rotor dyes offers a valuable and promising approach for streamlining the early formulation development process due to low material consumption and rapid analysis time in a 96-well plate format.