Procedures for Analytical Ultracentrifugation

超离心机是为数不多的能够提供有关溶液中大分子的分子量和相互作用的直接和可靠信息的仪器之一。沉降平衡技术以平衡热力学为基础，具有特殊的应用价值。它们提供了最可靠和万无一失的溶液参数估计，使用浓度分布的干涉观测具有极好的精度。沉降速度实验可以得到粒度/形状分布的详细信息。最近的进展使自动在线干涉测量数据采集成为一种常规和简单的程序。商业仪器目前在一种新的超离心机中提供高质量的吸光度测量。本研究将发展描述复杂大分子相互作用系统的程序和数据分析技术。平衡实验将主要通过各种最小二乘拟合程序进行分析，这将使溶质的物理化学参数在确定的置信度范围内进行广泛的全局拟合成为可能。将设计程序来精确和自动地监测平衡的实现。将探索平衡沉降实验的新方法。在平衡和速度实验中，将开发用于提高精密在线干涉测量的仪器。这些不同的方法将主要应用于生物技术企业感兴趣的问题，但不是全部。

The ultracentrifuge is one of the few instruments that provide direct and reliable information about molecular weights and interactions of macromolecules in solution. Sedimentation equilibrium techniques are especially valuable because of their foundation on equilibrium thermodynamics. They provide the most reliable and foolproof estimates of solution parameters with an excellent degree of precision using interferometric observations of concentration distributions. Sedimentation velocity experiments can yield detailed information about size/shape distributions. Recent advances have made automated on-line interferometric data acquisition a routine and simple procedure. Commercial instrumentation currently provides high-quality absorbance measurements in a new ultracentrifuge. This proposed research will develop procedures and data analysis techniques for characterizing difficult macromolecular interacting system. Equilibrium experiments will be analyzed primarily by various least squares fitting programs that will make possible extensive global fits to physicochemical parameters of the solute (s) with assured confidence limits. Programs will be devised for precisely and automatically monitoring attainment of equilibrium. Novel approaches to equilibrium sedimentation experiments will be explored. Instrumentation will be developed for significant improvement of precision on-line interferometry in both equilibrium and velocity experiments. These various approaches will be applied to problems of topical interest, largely, but not completely, from biotechnology enterprises.