Novel Hybrid Semiconductor-Cellular Standard for 3D FIB-SEM Nanotomography Analysis

Area of Science:

• Materials Science
• Microscopy
• Metrology

Background:

• Focused ion beam scanning electron microscopy (FIB-SEM) nanotomography offers high-resolution 3D imaging but relies heavily on instrument stability.
• Recognizing data disruptions is crucial for FIB-SEM's application as a metrology tool.

Purpose of the Study:

• To develop and evaluate a customizable standard sample for assessing FIB-SEM performance across different systems.
• To create a hybrid semiconductor-cellular standard for evaluating FIB-SEM acquisitions with biological samples.

Main Methods:

• Utilized a commercial semiconductor die with a uniform tungsten plug array for visualizing acquisition discontinuities.
• Cultured or drop-cast cells onto the die to create a hybrid standard.
• Quantified FIB-SEM dataset misalignment using affine transformation by comparing reconstructed and reference tungsten plug arrays.

Main Results:

• Abnormalities in tungsten plug position and morphology correlated with deviations in slice thickness.
• Misalignment quantification enables appropriate pixel data resizing and indicates potential recalibration needs.
• The developed standard sample is applicable to multiple FIB-SEM systems, enhancing nanotomography.

Conclusions:

• The proposed workflow provides a customizable, manufacturer-independent standard for improving FIB-SEM nanotomography.
• This method enhances the reliability and metrological capabilities of FIB-SEM by identifying and quantifying data acquisition issues.