Quantifying Cell Confluency by Plasmonic Nanodot Arrays to Achieve Cultivating Consistency

Abstract

The determination of cell confluency and subculture timing for cell culture consistency is crucial in the field of cell-based research, but there is no universal standard concerning optimal confluence. In this study, gold nanodot arrays on glass substrates were used as culture substrates, and their spectral shifts of localized surface plasmon resonance (LSPR) were employed to monitor cell growth and quantify cell confluency. Experiments including cell counting, metabolic activity, focal adhesion, and cell cycle were also performed to confirm the cell growth monitoring accuracy of the LSPR signals. The LSPR signal exhibited the same trends like the increase of cell numbers and cell metabolic activity and reached the maximum as the cell growth achieved confluency, suggesting its great capability as an effective indicator to predict suitable subculture timing. The proposed sensing approach is a noninterventional, nondestructive, real-time, and useful tool to help biologists quantify the optimal subculture timing, achieve cell culture consistency, and obtain reproducible experimental results efficiently.

Disposable Polymeric Nanostructured Plasmonic Biosensors for Cell Culture Adhesion Monitoring

“…Furthermore, Figures 2D,E show the plasmonic band displacement and the calibration curve, correspondingly; obtained by flowing different HCl solutions over the sensing areas, achieving a bulk refractive index sensitivity of 332.17 ± 0.71 nm/RIU in good agreement with the value previously reported by López-Muñoz (2021) . The obtained bulk sensitivity value is highly competitive and in the same order as those previously reported by engineered plasmonic nanostructures for cell adhesion monitoring ( Borile et al, 2019 ; Chang et al, 2019 ; Hou et al, 2019 ; Solis-Tinoco et al, 2019 ; Zhang et al, 2019 ). The LOD of the biosensing experimental set-up was calculated as previously reported ( López-Muñoz et al, 2017 ), the estimated value is near 3.43 × 10 −4 RIU (see Supplementary Figure S2 ).…”

“…Finally, the addition of an automated monitoring system could allow tracking adhesion kinetics with high time resolution (in the order of seconds) for long culture time (days or more) that could help to detect phenotypical changes in the cells. Aluminium nanopyramids (Zhang et al, 2019) Soft nanoimprint lithography Up to 475 nm/RIU Gold nanopillars (Solis-Tinoco et al, 2019) Shadow sphere lithography Up to 206 nm/RIU Gold nanodots (Chang et al, 2019) UV nanoimprint lithography ≈300 nm/RIU Gold nanogratings (Borile et al, 2019) Laser interference lithography 300 °/RIU Aluminium nanoslits (Hou et al, 2019) Thermal nanoimprint Up to 471 nm/RIU + Gold nanocrystals* Blu-ray discs (thermal nanoimprint) ≈330 nm/RIU + (Lee et al, 2017), *this article.…”

“…These advantages are mainly related to an enhanced surface sensitivity ( Špačková et al, 2018 ); and the potential to detect the surface interactions using simple optical arrays based on transmission/reflectance detection schemes instead of bulky and complex coupling methods (i.e., those based on attenuated total internal reflection by prism coupling) ( Lopez et al, 2017 ). There have been described different approaches of plasmonic biosensors based on nanostructured materials for cell monitoring in the last years ( Borile et al, 2019 ; Chang et al, 2019 ; Hou et al, 2019 ; Solis-Tinoco et al, 2019 ; Zhang et al, 2019 ); however, most of them involve a fabrication by complex lithographic methods and, as a result, lack of batch-to-batch reproducibility analysis of the plasmonic performance of the sensors.…”

Selective Growth of Patterned Monolayer Gold Nanoparticles on SU-8 through Photoreduction for Plasmonic Applications

“…Patterned metallic structures, which present potentially excellent properties on the nanometer scale, have attracted wide interest from the field of electronics, transparent conducting electrodes, supercapacitors, sensors, , polarization conversion, planar lens, color filter, perfect absorbers, hologram, surface-enhanced Raman spectroscopy, photovoltaics, photothermal therapy, and biomedical applications. , For a wide variety of applications, a reliable process that can easily control the sizes, geometries, and spatial distributions of metallic nanostructures is necessary. Conventionally, nanofeatures are patterned through electron beam lithography (EBL) or optical lithography followed by metal deposition and lift-off processes.…”

A novel method of cell culture based on the microfluidic chip for regulation of cell density

“…However, feedback depends on human manipulation, which is laborious and highly subjective (Wu et al, 2016). The feedback regulation may cause the differences of cell density in cell culture, ultimately resulting in the inaccuracy of the repeatability of dose‐response assays in anticancer drug development (Chang et al, 2019). Thus, it has become increasingly important to develop a precise cell culture method with the automatic control of cell density to achieve density consistency.…”