Quantitative phase microscopies: accuracy comparison

Area of Science:

• Biophysics
• Optical Microscopy
• Quantitative Imaging

Background:

• Quantitative phase microscopies (QPMs) are crucial for bio-imaging, providing label-free data on mass distribution and transport.
• QPMs complement fluorescence imaging by avoiding photobleaching and phototoxicity.
• Selecting the optimal QPM technique for specific applications can be challenging due to the variety available.

Purpose of the Study:

• To provide a comprehensive tutorial review comparing the main QPM techniques.
• To focus on the accuracy of QPMs, specifically measurement precision and trueness.
• To assist researchers in choosing the most suitable QPM for their needs.

Main Methods:

• Comparison of 8 QPM techniques: digital holographic microscopy (DHM), cross-grating wavefront microscopy (CGM), diffraction phase microscopy (DPM), differential phase-contrast (DPC) microscopy, phase-shifting interferometry (PSI) imaging, Fourier phase microscopy (FPM), spatial light interference microscopy (SLIM), and transport-of-intensity equation (TIE) imaging.
• Utilized a custom numerical toolbox based on discrete dipole approximation (IF-DDA) to model electromagnetic fields.
• Upgraded the toolbox to simulate various QPMs and incorporate shot noise effects.

Main Results:

• Digital holographic microscopy (DHM) and phase-shifting interferometry (PSI) are largely artifact-free, primarily affected by coherent noise.
• Cross-grating wavefront microscopy (CGM), differential phase-contrast (DPC) microscopy, diffraction phase microscopy (DPM), and transport-of-intensity equation (TIE) imaging show a precision-trueness trade-off adjustable via experimental parameters.
• Fourier phase microscopy (FPM) and spatial light interference microscopy (SLIM) exhibit inherent artifacts, limiting their quantitative accuracy, especially for large biological specimens like eukaryotic cells.

Conclusions:

• The study provides a quantitative comparison of major QPM techniques based on precision and trueness.
• Results guide the selection of QPMs by highlighting their strengths and limitations regarding accuracy and artifacts.
• The developed numerical toolbox facilitates the modeling and analysis of QPM performance.