Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Two-Step Thermal Transformation of Multilayer Graphene Using Polymeric Carbon Source Assisted by Physical Vapor Deposited Copper

Direct in situ growth of graphene on dielectric substrates is a reliable method for overcoming the challenges of complex physical transfer operations, graphene performance degradation, and compatibility with graphene-based semiconductor devices. A transfer-free graphene synthesis based on a controllable and low-cost polymeric carbon source is a promising approach for achieving this process. In this paper, we report a two-step thermal transformation method for the copper-assisted synthesis of transfer-free multilayer graphene. Firstly, we obtained high-quality polymethyl methacrylate (PMMA) film on a 300 nm SiO2/Si substrate using a well-established spin-coating process. The complete thermal decomposition loss of PMMA film was effectively avoided by introducing a copper clad layer. After the first thermal transformation process, flat, clean, and high-quality amorphous carbon films were obtained. Next, the in situ obtained amorphous carbon layer underwent a second copper sputtering and thermal transformation process, which resulted in the formation of a final, large-sized, and highly uniform transfer-free multilayer graphene film on the surface of the dielectric substrate. Multi-scale characterization results show that the specimens underwent different microstructural evolution processes based on different mechanisms during the two thermal transformations. The two-step thermal transformation method is compatible with the current semiconductor process and introduces a low-cost and structurally controllable polymeric carbon source into the production of transfer-free graphene. The catalytic protection of the copper layer provides a new direction for accelerating the application of graphene in the field of direct integration of semiconductor devices