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Implement Structure Imaging Page (#57)
* Added initial view * Add content * Prettified Code! * Test deploy --------- Co-authored-by: daniel-panhead <[email protected]>
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<script setup lang="ts"> | ||
import Notebook from '@/components/lab-notebook/Notebook.vue' | ||
import SingleColumn from '@/components/lab-notebook/SingleColumn.vue' | ||
import CaptionedGraphics from '@/components/CaptionedGraphics.vue' | ||
const sectionTitleStyle = 'text-subtitle-sm lg:text-subtitle text-white mb-4' | ||
</script> | ||
|
||
<template> | ||
<Notebook> | ||
<template #title> Structure Imaging </template> | ||
<template #body> | ||
<SingleColumn> | ||
<template #title> Overview </template> | ||
<template #body> | ||
<p> | ||
To visualize the formation of the octahedron, Atomic Force Microscopy (AFM) was employed | ||
due to its capacity for high-resolution imaging and its established utility in | ||
visualizing DNA origami structures. An attempt was made to observe octahedron formation; | ||
however, this endeavor was impeded by a machine calibration malfunction, preventing a | ||
conclusive determination of the success of octahedron formation. Although the acquired | ||
images suggest a probable formation of the octahedron structure, future attempts are | ||
imperative, using the same methods outlined below, in order to definitively confirm its | ||
formation. | ||
</p> | ||
</template> | ||
</SingleColumn> | ||
|
||
<SingleColumn> | ||
<template #title> Background </template> | ||
<template #body> | ||
<p> | ||
Atomic Force Microscopy (AFM) is a technique used to obtain high-resolution images of | ||
nanoscale structures, including DNA origami and protein assemblies (Kolbeck et al., | ||
2023). AFM employs a sharp probe to scan the surface of the sample mechanically. The | ||
interaction between the probe tip and the sample surface generates a topographical/3D | ||
map, which can be magnified for detailed examination(Kolbeck et al., 2023). This allows | ||
researchers to visualize intricate structures at the nanometer scale without the need | ||
for electron beams (Kolbeck et al., 2023). AFM is particularly useful for studying | ||
biological samples, as it does not require a vacuum environment or special sample | ||
preparation, making it more versatile for a wide range of applications (Kolbeck et al., | ||
2023). | ||
</p> | ||
</template> | ||
</SingleColumn> | ||
|
||
<SingleColumn> | ||
<template #title> Aims </template> | ||
<template #body> | ||
<ol class="list-decimal pl-8"> | ||
<li> | ||
To observe and confirm the synthesis of DNA origami structures, to serve as a scaffold | ||
for imaging | ||
</li> | ||
<li>To determine the extent of chaining/polymerization for octahedrons</li> | ||
<li>To demonstrate the viability of using AFM for DNA origami methods</li> | ||
</ol> | ||
</template> | ||
</SingleColumn> | ||
|
||
<SingleColumn> | ||
<template #title> Methods </template> | ||
<template #body> | ||
<div class="flex flex-col gap-y-4"> | ||
<p> | ||
The method was adapted by Lv et al., 2015. To capture detailed AFM (Atomic Force | ||
Microscopy) images of DNA nanostructures, a mica surface was employed as the | ||
substrate. Mica is a silicate mineral that is frequently used in microscopy due to its | ||
smooth, reactive surface. Initially, the mica surface was prepared by cleaving it with | ||
a sterilized, sharp scalpel, although Scotch tape can also be employed for this task. | ||
Following the cleavage, we applied a prepared buffer onto the mica and let it incubate | ||
for 5 minutes. To ensure complete removal of the buffer, the surface was washed with 1 | ||
mL of double-distilled water (ddH2O). This washing step was conducted three times. | ||
</p> | ||
<p> | ||
Subsequently, 2.5µL of previously prepared DNA nanostructures were applied onto the | ||
mica. The sample was then allowed to incubate for 5 minutes, ensuring that the DNA | ||
would securely adhere to the mica surface for stable imaging. To minimize the risk of | ||
salt contamination, we rinsed the mica surface with 1 mL of either Millipore water or | ||
ddH2O. After washing, the mica surface was immediately dried under Argon for 1 minute. | ||
</p> | ||
<p> | ||
Finally, with both the mica surface and DNA nanostructures properly prepared, the | ||
sample was ready for AFM imaging. | ||
</p> | ||
</div> | ||
</template> | ||
</SingleColumn> | ||
|
||
<SingleColumn> | ||
<template #title> Results </template> | ||
<template #body> | ||
<div class="flex flex-col gap-y-4"> | ||
<CaptionedGraphics> | ||
<template #graphics> | ||
<div class="grid md:grid-cols-2 grid-rows-2 gap-10 lg:w-3/4"> | ||
<img src="../../assets/structure-imaging/sample-1.jpeg" alt="" class="h-full" /> | ||
<img src="../../assets/structure-imaging/sample-2.jpeg" alt="" class="h-full" /> | ||
<img src="../../assets/structure-imaging/sample-3.jpeg" alt="" class="h-full" /> | ||
<img src="../../assets/structure-imaging/sample-4.jpeg" alt="" class="h-full" /> | ||
</div> | ||
</template> | ||
<template #caption> | ||
<p> | ||
Figure 1. Four AFM images of the area of Octahedron sample (25 nM). The X axis | ||
represents width and the Y axis represents length of the samples. The range of | ||
values on the sidebars represent the heights of samples (length in the Z | ||
dimension). It should be noted that these values are not reliable due to | ||
calibration errors of the machine. | ||
</p> | ||
</template> | ||
</CaptionedGraphics> | ||
<p> | ||
In the acquired images, we observed distinct white "blobs" that may potentially | ||
represent octahedron structures. However, it is important to note that the reliability | ||
of the xyz axes is compromised due to a calibration malfunction in the imaging | ||
machine. As a result, we are unable to provide conclusive information regarding the | ||
sizes of these structures, even when using a standard reference sample. | ||
</p> | ||
</div> | ||
</template> | ||
</SingleColumn> | ||
|
||
<SingleColumn> | ||
<template #title> Discussion </template> | ||
<template #body> | ||
<div class="flex flex-col gap-y-6"> | ||
<p> | ||
While the features observed in the AFM images resemble octahedrons, they could also | ||
potentially be salt crystals, specifically MgCl2, which is commonly used in the | ||
thermoramp protocol for forming octahedrons from scaffold and staple strands. However, | ||
it's important to note that the absence of an accurate scale bar makes it | ||
inappropriate to draw conclusions without further verification. Therefore, retesting | ||
AFM or employing an alternative validation method is necessary. The gel | ||
electrophoresis results did exhibit a noticeable band shift for the formed structure | ||
compared to the scaffold. Nevertheless, it's worth noting that gel electrophoresis | ||
provides more of a qualitative assessment, and for greater confidence in the | ||
successful formation of octahedrons, we need to complement it with AFM or other | ||
visualization techniques. In our future experiments, we plan to utilize AFM in | ||
conjunction with TEM (Transmission Electron Microscopy) to establish two reliable | ||
methods for confirming the structure's formation. AFM will be our initial choice due | ||
to its quicker results. However, it's essential to recognize that AFM primarily | ||
confirms the DNA aspects of the octahedron. For a comprehensive understanding of | ||
liposome formation templated on the octahedron, cryo-TEM remains the most suitable | ||
method. | ||
</p> | ||
</div> | ||
</template> | ||
</SingleColumn> | ||
|
||
<SingleColumn :always-dropdown="true"> | ||
<template #title> References </template> | ||
<template #body> | ||
<div class="pl-6 -indent-6"> | ||
<p> | ||
Kolbeck, P. J., Dass, M., Martynenko, I. V., van Dijk-Moes, R. J. A., Brouwer, K. J. | ||
H., van Blaaderen, A., Vanderlinden, W., Liedl, T., & Lipfert, J. (2023). DNA Origami | ||
Fiducial for Accurate 3D Atomic Force Microscopy Imaging. Nano Letters, 23(4), | ||
1236–1243. https://doi.org/10.1021/acs.nanolett.2c04299 | ||
</p> | ||
<p> | ||
Lv, Y., Hu, R., Zhu, G., Zhang, X., Mei, L., Liu, Q., Qiu, L., Wu, C., & Tan, W. | ||
(2015). Preparation and biomedical applications of programmable and multifunctional | ||
DNA nanoflowers. Nature Protocols, 10(10), 1508–1524. | ||
https://doi.org/10.1038/nprot.2015.078 | ||
</p> | ||
</div> | ||
</template> | ||
</SingleColumn> | ||
</template> | ||
</Notebook> | ||
</template> |