Atomic Force Microscopy

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Atomic force microscopy (AFM) has emerged as a powerful characterization platform, providing valuable insight that may have major implications for drug design and development. Creative Biolabs has extensive project experience in mRNA delivery research. We offer custom solutions with our cutting-edge facility, knowledge, and technology. Here are some examples of AFM technology in use.

Chemical and Structural Investigation of Lipid Nanoparticles (LN) by AFM

AFM has recently been employed as a major microscopic tool to observe the size and morphology of LN and nanoemulsions. Due to the soft nature of all samples, AFM with a tapping mode was used in this study to characterize the molecular structure at varying liquid oil loading. In this mode, an oscillating probe tip taps, at high frequency, the samples while scanning them. The advantages of this mode are that the low forces and minimal damage enable it to image soft samples in the air. Discrete and disc-like shapes were found in Fig.1. No significant difference in particulate morphology between lipid nanoparticles at varying concentrations of liquid lipid was observed, but the differences in particulate phase images were found. The phase separation of nanoparticles was observed with the 0% liquid lipid system. But this became less apparent for the 5% and 10% liquid lipid systems.

The size of the LN was also measured from the AFM images. The geometric mean diameters of 0%, 5%, and10% liquid lipid were 100.5±28.7 nm, 91.2±16.8 nm, and 85.8±23.1 nm, respectively, whereas the mean heights were found to be 9.1±3.1 nm, 5.7±0.6 nm, and 3.1±0.7 nm, respectively. Moreover, there is a large discrepancy in the obtained size from AFM and PCS, which could be due to the method of preparation. For the AFM measurement, material dehydration during sample preparation may occur, whereas for the PCS measurement the particles were suspended in an aqueous medium or a swelling state. Therefore, smaller particle sizes from AFM or TEM measurements as compared to PCS have been obtained and previously reported. In conclusion, AFM revealed the phase-separated structure particles for the pure solid lipid system, whereas it was less apparent in the presence of the liquid lipid. These findings should be beneficial to future designs of nanoparticles as drug delivery systems.

AFM morphology of γ-oryzanol loaded LNs at 0% liquid lipid (A), 5% liquid lipid (B), and 10% liquid lipid (C): 3D images (1), topographic images (2) and phase images (3). All images were scanned over a 1×μm2 area.Fig.1 AFM morphology of γ-oryzanol loaded LNs at 0% liquid lipid (A), 5% liquid lipid (B), and 10% liquid lipid (C): 3D images (1), topographic images (2), and phase images (3). All images were scanned over a 1×μm2 area. (Anantachaisilp, 2010)

The Ultrastructure of Collagen/Lipid Nanoparticle Cryostructurates was Explored by AFM

Collagen cryo structures appeared comprising filamentous and laminar structures made of compact fibrils exhibiting the typical banding patterns of native collagens. When observed by AFM, nanoparticles appeared as facetted nanostructures in the form of aggregates or attached to collagen fibers and were slightly larger than lipid particles. Interestingly, the D-band spacing pattern of collagen/LNP samples was not significantly modified, indicating that collagen fibers were well preserved in the presence of nanoparticles. The overall structural organization of these collagen fibers could be maintained for nanoparticle concentration up to a collagen/lipid ratio of 2/1, while it became more relaxed for collagen/lipid ratios below 2.5/1.

AFM analysis (in the air) of collagen (C) or collagen/lipid nanoparticle films (A, B: I-images; II-particle size analysis). D) Band spacing analysis of the collagen fibers: I-height profile along the arrows indicated in images (AI, BI, C); II-mean band spacing measured for different samples.Fig.2 AFM analysis (in the air) of collagen (C) or collagen/lipid nanoparticle films (A, B: I-images; II-particle size analysis). D) Band spacing analysis of the collagen fibers: I-height profile along the arrows indicated in images (AI, BI, C); II-mean band spacing measured for different samples. (Laghezza, 2019)

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References

  1. Anantachaisilp, S.; et al. Chemical and structural investigation of lipid nanoparticles: drug-lipid interaction and molecular distribution. Nanotechnology. 2010, 21(12): 125102.
  2. Laghezza, M.V.; et al. Characterization of Collagen/Lipid Nanoparticle-Curcumin Cryostructurates for Wound Healing Applications. Macromolecular bioscience. 2019, 19(5): 1800446.
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