19y and 62y old NHDFs were seeded in an 8-well tissue culture treated glass slide (Falcon, Cat# 354118) at 20,000 cells/well. Cells were incubated at standard conditions in cell culture media. After 24 h, cells were treated with 2% vol/vol of Adansonia digitata in DMEM or a media control for 48 h, with treatment refreshed at 24 h. Following treatment, miR-146a was labeled in situ using the ViewRNA Cell Plus Assay from ThermoFisher (Cat#88-19000-99), as per manufacturers’ protocol, with the appropriate target probe set for miR-146a (Cat#VM1-10253-06). To summarize, cells were first fixed and permeabilized. Next, cells were incubated in a blocking solution. Cells were then incubated with a solution of the miR-146a targeting probe, followed subsequently by solutions with PreAmplifier probes, Amplifier probes, and finally Type 1 Label probes that are conjugated to the Alexa Fluor 546 synthetic dye. Cell nuclei were then stained using a DAPI solution. Cells were mounted in Prolong Glass mounting medium (ThermoFisher, Cat#P36982) with a No. 1.5 cover slip (Corning, Cat#2980-245), and allowed to cure for at least 24 h at RT prior to image acquisition. All images were captured on a Nikon A1-HD 25 laser scanning confocal microscope using a 60×, 1.4 NA oil immersion objective.
No 1.5 coverslip
Manufactured by Corning
Sourced in United States
The No. 1.5 coverslip is a thin, transparent glass or plastic slide used in microscopy applications. It is designed to be placed over a specimen on a microscope slide, providing a protective barrier and facilitating the imaging process.
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3 protocols using no 1.5 coverslip
1
In Situ miR-146a Quantification in NHDFs
2
Single Particle Studies of AuNRs@mSiO2
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The samples for single particles studies were prepared as follows: the colloid solution was first diluted with 18.2 MΩ pure water to a proper concentration. The diluted solution was then sonicated for 10 min at room temperature. Samples were prepared by drop casting the diluted AuNRs@mSiO2 solution on pre-cleaned glass slides. Subsequently, a 22 mm × 22 mm No. 1.5 coverslip (Corning, NY, USA) was placed on the glass slide. Throughout all of the experiments, the concentration of AuNRs@mSiO2 deposited on the glass slide surface was maintained at about 1 μm−2 to facilitate single particle characterization without inter-particle interaction and LSPR coupling.
3
Characterization of Pt-Coated Gold Nanorods
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Bare AuNRs (25 × 73 nm) without other metals on the surface were obtained from Nanopartz (Loveland, CO, USA) (Fig. S1), and Pt-AuNRs with an average size of 25 × 73 nm were purchased from Sigma-Aldrich (St. Louis, MO, USA). The Pt-AuNR colloid solution was first diluted with 18.2-MΩ pure water to a proper concentration. The diluted solution was then sonicated for 15 min at room temperature. Samples were prepared by spin casting the solution on the pre-cleaned glass slide. Then, a 22 × 22 mm No. 1.5 coverslip (Corning, NY) was covered on the glass slide. In this study, the concentration of Pt-AuNRs on the glass surface was controlled to be ~1 μm -2 in order to facilitate single-particle characterization and to minimize inter-particle SPR coupling, resulting in a spectral shift. Structural characterization was carried out using a transmission electron microscope (TEM) (H-8100, Hitachi, Japan).
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