Darkfield Microscopy to Visualize Diffusional Dynamics of Nanorods on Cell Membrane
Published: July 31, 2023
Abstract
Source: Ge, F., et al. Visualizing Diffusional Dynamics of Gold Nanorods on Cell Membrane using Single Nanoparticle Darkfield Microscopy. J. Vis. Exp. (2021).
In this video, we describe the darkfield microscopy (DFM) technique combined with single-particle tracking (SPT) analysis to monitor the interaction of plasmonic gold nanorods with the cell membrane of a specific cell line of interest. Combined DFM and SPT analysis allows the visualization of the nanorod movement through the cell membrane with high spatiotemporal resolution.
Protocol
1. Microscope slide preparation NOTE: U87 MG cells of the third to tenth generation with high activity are used in SPT experiments. Sterilize 22 mm × 22 mm coverslips already cleaned with Piranha solution by immersing in ethanol (99.9%). Use forceps to take out the cover slip from the ethanol solution (step 1.1) and sterilize by burning ethanol on the flame. Once all ethanol is burnt, place coverslips in a plastic cell culture dish (…
Representative Results
Figure 1: Schematic diagram of the optical path of darkfield microscopy, microscope slide preparation, and the dual-channel difference method for calculating polar angle (θ) of AuNRs. Scale bar in image is 2 μm. Figure 2: Characterization of CTAB-AuNRs. (A) TEM images of CTAB@AuNRs….
Disclosures
The authors have nothing to disclose.
Materials
| CTAB-coated gold nanorods (CTAB-AuNRs) | Nanoseedz | NR-40-650 | 85 nm*40 nm |
| Color CMOS camera | Olympus | DP74 | Japan |
| Coverslips | Citoglas | z10212222C | 22*22 mm |
| Dark-field microscopy | Nikon | 80i | Upright microscope |
| Fetal bovine serum (FBS) | Gibco | 10099141 | |
| Fiji | National Institutes of Health | 2.0.0-rc-69/1.52 p | A distribution of ImageJ |
| Grooved glass slide | Sail brand | 7103 | Single concave |
| Image J | Image J | 1.52 j | |
| MATLAB | MathWorks | R2019b | |
| MATLAB Code | https://github.com/fenggeqd/ JOVE-2020 | ||
| Plastic cell culture dishes | Falcon | 353002 | |
| Plastic cell culture dishes | Falcon | 353001 | 35*10 mm |
| U87 MG cell | American Type Culture Collection | ATCC HTB-14 | A human primary glioblastoma cell line |
| Minimum essential medium (MEM) | Gibco | 51200038 | No phenol red |
| Penicillin-streptomycin | Gibco | 15140122 |
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Cite This Article
Darkfield Microscopy to Visualize Diffusional Dynamics of Nanorods on Cell Membrane. J. Vis. Exp. (Pending Publication), e21458, doi: (2023).