<p>Quantitatively, none of the LNP-treated samples showed significant fluorescence, indicating a failure in transfection. The lack of fluorescence in all experimental groups, except the positive control, suggests either insufficient uptake of the LNPs by the cells or a failure in expression of the YFP reporter. </p>
<p>Quantitatively, none of the LNP-treated samples showed significant fluorescence, indicating a failure in transfection. The lack of fluorescence in all experimental groups, except the positive control, suggests either insufficient uptake of the LNPs by the cells or a failure in expression of the YFP reporter. </p>
<p>Fluorescence microscopy with the Leica DMI6000 B microscope at 20x magnification was by performed us on HEK293 cells transfected with LNPs containing pcDNA 3.1 eYFP DNA and mRNA. Minicircle DNA served as the positive control, while LNPs without cargo acted as the negative control. Cells were imaged at 24 h, 48 h, and 72 h post-transfection.</p>
<p>Quantitatively, after 72 h none of the LNP-treated samples showed significant fluorescence, indicating a failure in transfection (Figure 21). The lack of fluorescence in all experimental groups, except lipofectamine and Minicircle DNA as the positive control, suggests either insufficient uptake of the LNPs by the cells or a failure in expression of the YFP reporter, indicating that the Corden LNP may not suited as our delivery system. Also the deformed morphology and decreased growth are indicators for negative effects of the Cayman LNP on HEK293, probably reasoned in the employment of more cytotoxic mPEG-DSPE compared to DMG-PEG in the Cayman and SORT LNP.</p>
Fluorescence microscopic images of transfected HEK293 cells at 20x magnification after 48 h post-transfection with different Corden LNP formulations recorded with Leica DMI6000 B.
Overlay of phase contrast and fluorescence microscopic images of transfected HEK293 cells at 20x magnification after 72 h post-transfection with different Corden LNP formulations recorded with Leica DMI6000 B.
</figcaption>
</figcaption>
</figure>
</figure>
<H5text="Cryo-EM"/>
<H5text="Cryo-EM"/>
<p>Cryo-EM (cryogenic electron microscopy) as a form of transmission electron microscopy (TEM) was performed by us using a JEOL JEM-2200FS electron microscope (JEOL, Freising, Germany) operating at 200kV, equipped with a cold field emission electron gun. The sample preparation and imaging were carried out at cryogenic temperatures, which allowed for the visualization of LNPs in their native hydrated state.</p>
<p>Cryo-EM (cryogenic electron microscopy) as a form of transmission electron microscopy (TEM) was performed by us using a JEOL JEM-2200FS electron microscope (JEOL, Freising, Germany) operating at 200kV, equipped with a cold field emission electron gun. The sample preparation and imaging were carried out at cryogenic temperatures, which allowed for the visualization of LNPs in their native hydrated state.</p>
