A radial calibration window for analytical ultracentrifugation

Area of Science:

• Biophysical chemistry
• Analytical chemistry
• Materials science

Background:

• Analytical ultracentrifugation (AUC) is a powerful technique for characterizing macromolecules and particles in solution.
• Previous studies identified radial dimension calibration accuracy as a major limitation in AUC measurements.
• Significant errors (1-15%) in radial measurements were observed across various AUC instruments.

Purpose of the Study:

• To develop a novel artifact for accurate, absolute calibration of radial magnification in AUC.
• To enhance the precision and reliability of AUC measurements for hydrodynamic and particle size analysis.

Main Methods:

• Development of a lithographically deposited reflective pattern artifact on an AUC window.
• Utilizing the artifact as a reticle for scanning in AUC control experiments.
• Analysis of landmark pitch in scans from different optical systems to determine radial magnification accuracy.

Main Results:

• The developed artifact enables absolute calibration of radial magnification in AUC.
• Residual uncertainty in radial magnification after calibration is estimated at approximately 0.2%.
• This artifact can reduce radial measurement errors by 5- to 10-fold or more compared to previous methods.

Conclusions:

• External radial calibration using the developed artifact is crucial for improving AUC accuracy.
• Adoption of this artifact will enhance the reliability of molecular hydrodynamics and particle size measurements.
• This innovation addresses a key limitation, leading to more precise and trustworthy AUC data.