Improved measurement of the rotor temperature in analytical ultracentrifugation

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• 作者：Huaying Zhao^a ; Andrea Balbo^b ; Howard Metger^c ; Robert Clary^c ; Rodolfo Ghirlando^d ; ^{rodolfo.ghirlando@nih.gov" class="auth_mail} ; Peter Schuck^a ; ^{schuckp@mail.nih.gov" class="auth_mail}
• 关键词：Sedimentation velocity ; Temperature calibration
• 刊名：Analytical Biochemistry
• 出版年：15 April, 2014
• 年：2014
• 卷：451
• 期：Complete
• 页码：69-75
• 全文大小：1263 K

文摘

Sedimentation velocity is a classical method for measuring the hydrodynamic, translational friction coefficient of biological macromolecules. In a recent study comparing various analytical ultracentrifuges, we showed that external calibration of the scan time, radial magnification, and temperature is critically important for accurate measurements (Anal. Biochem. 440 (2013) 81-95). To achieve accurate temperature calibration, we introduced the use of an autonomous miniature temperature logging integrated circuit (Maxim Thermochron iButton) that can be inserted into an ultracentrifugation cell assembly and spun at low rotor speeds. In the current work, we developed an improved holder for the temperature sensor located in the rotor handle. This has the advantage of not reducing the rotor capacity and allowing for a direct temperature measurement of the spinning rotor during high-speed sedimentation velocity experiments up to 60,000 rpm. We demonstrated the sensitivity of this approach by monitoring the adiabatic cooling due to rotor stretching during rotor acceleration and the reverse process on rotor deceleration. Based on this, we developed a procedure to approximate isothermal rotor acceleration for better temperature control.