Development of Adaptive SEM Technology for Genome/Proteome Expression Analysis in Single Cell Level

Abstract

Identification of each cellular phenotype in a tissue is essential for understanding community effect in living system. One useful way is in situ measurement of expressed biomarkers in single cell level using a lot labels; however, production and identification of such labels are still challenging. We propose a new sensing technology, which is a comprehensive development of production and identification of nano-particle (NP) labels for simultaneous in situ measurements of expressed biomarkers in single cell.For the fabrication of NPs, various sizes of polystyrene spheres were used as casts, and metals were deposited on the spheres by thermal evaporation. When polystyrene casts were interfering with measurements, degradation of organic substances was performed, and cup-shaped metal NP shells can be also obtained. For the identification of fabricated NPs, field emission scanning electron microscopy (FE-SEM) was used. Both diameters and elements of NPs were identified with observations of secondary electron (SE) and backscattered electron (BE) in the FE-SEM.By using our method, more than 500 types of NPs were fabricated. Metal shell layers were formed by thermal evaporation; therefore, multi-layered NPs can be easily fabricated with sequential evaporation. We used double-layered NPs; outer is Au for easy immobilization of biomolecules to use these NPs as labels of biomarkers, and inner layer is various to apply label varieties. Spatial distributions and diameters of NP labels were identified by SE observation, and NP elements were identified by BE observation as the difference of intensities in the image caused by the difference of atomic number of inner metal. We call it “adaptive SEM” technology (i.e., NP identification is adaptive for various samples). These results indicate a possibility for quantitative in situ detection of expressed biomarkers in a cell by the suggested technology.