Lambda Theta Reflectometry: A New Technique for Measuring Optical Film Thickness in Planar Protein Arrays

Quantitative protein measurements provide valuable information about biological pathways, immune system functionality, and the mechanisms of disease. The most accurate methods for detecting proteins are label-free and preserve native protein-binding interactions. Label-free biomolecular interaction analysis includes reflectometry, a group of techniques that detect proteins by measuring the reflectance properties of a thin film on a substrate. Most of these techniques are limited in some way by instrument complexity, sensitivity, or consumable manufacturing requirements. To address these issues, we introduce Lambda Theta Reflectometry (LTR), a new reflectometric technique that measures changes in film thickness by determining the point of null reflectivity as a function of wavelength (lambda) and angle of incidence (theta). The substrate is simultaneously illuminated with a range of angles and wavelengths, and reflected light is resolved both angularly and spectrally. Our prototype LTR reflectometer can measure SiO₂ layer thickness with milli-Ångström precision. LTR measurements of Si/SiO₂ oxide films are in excellent agreement with spectroscopic ellipsometry for film thicknesses ranging from 1390 to 1465 Å. This technique enables label-free biosensing measurements across a range of biological analyte concentrations (0.5 ng/mL to μg/mL) without requiring stringent control over probe deposition thickness or substrate manufacturing.