Contribution of metal layer thickness for quantitative backscattered electron imaging of field emission scanning electron microscopy

The contributions of metal thickness and the diameter of metal shell particles to quantitative backscattered electron (BSE) imaging in field emission scanning electron microscopy (FE-SEM) were studied to evaluate the potential of using these particles as simultaneously distinguishable labels of target molecules in FE-SEM studies. Gold spherical shells were fabricated with 200 or 300 nm diameter and 5, 10, 15 or 20 nm Au shell thickness, placed on silicon substrates, respectively, and observed in BSE imaging mode by FE-SEM. Flat Au films of the same thicknesses were formed on a Si substrate to evaluate the contribution of shell diameter to BSE imaging. The relationship between relative BSE intensity, which was calculated by setting the intensities of the Si substrate and 20 nm-thick Au layer as standards, and Au layer thickness was studied for all samples. With increasing Au layer thickness, BSE intensity also proportionally increased for all samples (R² > 0.93) in the range of these thicknesses. Gradients of the increase were 1.5 times different between the flat film and metal shells, which was caused by the presence of voids in shell particles. The difference in gradients for increasing shell thickness between 200 and 300 nm particles was 15%. This result indicated that 1.5 times difference in shell diameter contributed to the increase of BSE intensity against the increase of shell thicknesses as a 15% error, and strict control of both metal shell diameter and thickness in the fabrication process is essential when using these shells as labels of BSE measurements in FE-SEM.