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Biological Atomic Force Microscopy for Imaging Gold-Labeled Liposomes on Human Coronary Artery Endothelial Cells.

Zaske AM, Danila D, Queen MC, Golunski E, Conyers JL - J Pharm (Cairo) (2013)

Bottom Line: Dynasore monohydrate, an inhibitor of endocytosis, obstructed the internalization of the gold-liposomes.This study showed the versatility of the AFM technique, combined with fluorescent microscopy, for investigating liposome uptake by endothelial cells.The 90 nm colloidal gold nanoparticles proved to be a noninvasive contrast agent that efficiently improves AFM imaging during the investigation of biological nanoprocesses.

View Article: PubMed Central - PubMed

Affiliation: Division of Cardiology, Department of Internal Medicine, The University of Texas Health Science Center at Houston, 1881 East Road, Houston, TX 77054, USA.

ABSTRACT
Although atomic force microscopy (AFM) has been used extensively to characterize cell membrane structure and cellular processes such as endocytosis and exocytosis, the corrugated surface of the cell membrane hinders the visualization of extracellular entities, such as liposomes, that may interact with the cell. To overcome this barrier, we used 90 nm nanogold particles to label FITC liposomes and monitor their endocytosis on human coronary artery endothelial cells (HCAECs) in vitro. We were able to study the internalization process of gold-coupled liposomes on endothelial cells, by using AFM. We found that the gold-liposomes attached to the HCAEC cell membrane during the first 15-30 min of incubation, liposome cell internalization occurred from 30 to 60 min, and most of the gold-labeled liposomes had invaginated after 2 hr of incubation. Liposomal uptake took place most commonly at the periphery of the nuclear zone. Dynasore monohydrate, an inhibitor of endocytosis, obstructed the internalization of the gold-liposomes. This study showed the versatility of the AFM technique, combined with fluorescent microscopy, for investigating liposome uptake by endothelial cells. The 90 nm colloidal gold nanoparticles proved to be a noninvasive contrast agent that efficiently improves AFM imaging during the investigation of biological nanoprocesses.

No MeSH data available.


Related in: MedlinePlus

AFM images of non-gold-coupled liposomes visualized in amplitude (a) and height (b) modes to 1.7 μm (x-y). The AFM section analysis of these liposomes showed an average diameter of 121.5 ± 27 nm when deposited on mica and scanned in air. This result was comparable to that of the DLS analysis (129 ± 1.3 nm). Scan obtained in tapping mode using RTESP tips (fo = 262–325 kHz, k = 20–80 N/m).
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fig2: AFM images of non-gold-coupled liposomes visualized in amplitude (a) and height (b) modes to 1.7 μm (x-y). The AFM section analysis of these liposomes showed an average diameter of 121.5 ± 27 nm when deposited on mica and scanned in air. This result was comparable to that of the DLS analysis (129 ± 1.3 nm). Scan obtained in tapping mode using RTESP tips (fo = 262–325 kHz, k = 20–80 N/m).

Mentions: The structural properties of uncoupled and gold-coupled liposomes were analyzed using AFM. Both types of liposomes were imaged after fixation with 10% formalin to ensure the preservation of their original structure. AFM was performed in tapping mode in air with RTESP cantilevers. Figure 2 shows a typical AFM image of non-gold-coupled liposomes deposited on fresh cleaved mica and fixed with formalin. AFM analysis of these liposomes showed an average diameter of 121.5 ± 27 nm. This result was comparable to the diameter measurement obtained by DLS (129 ± 1.3 nm).


Biological Atomic Force Microscopy for Imaging Gold-Labeled Liposomes on Human Coronary Artery Endothelial Cells.

Zaske AM, Danila D, Queen MC, Golunski E, Conyers JL - J Pharm (Cairo) (2013)

AFM images of non-gold-coupled liposomes visualized in amplitude (a) and height (b) modes to 1.7 μm (x-y). The AFM section analysis of these liposomes showed an average diameter of 121.5 ± 27 nm when deposited on mica and scanned in air. This result was comparable to that of the DLS analysis (129 ± 1.3 nm). Scan obtained in tapping mode using RTESP tips (fo = 262–325 kHz, k = 20–80 N/m).
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4590807&req=5

fig2: AFM images of non-gold-coupled liposomes visualized in amplitude (a) and height (b) modes to 1.7 μm (x-y). The AFM section analysis of these liposomes showed an average diameter of 121.5 ± 27 nm when deposited on mica and scanned in air. This result was comparable to that of the DLS analysis (129 ± 1.3 nm). Scan obtained in tapping mode using RTESP tips (fo = 262–325 kHz, k = 20–80 N/m).
Mentions: The structural properties of uncoupled and gold-coupled liposomes were analyzed using AFM. Both types of liposomes were imaged after fixation with 10% formalin to ensure the preservation of their original structure. AFM was performed in tapping mode in air with RTESP cantilevers. Figure 2 shows a typical AFM image of non-gold-coupled liposomes deposited on fresh cleaved mica and fixed with formalin. AFM analysis of these liposomes showed an average diameter of 121.5 ± 27 nm. This result was comparable to the diameter measurement obtained by DLS (129 ± 1.3 nm).

Bottom Line: Dynasore monohydrate, an inhibitor of endocytosis, obstructed the internalization of the gold-liposomes.This study showed the versatility of the AFM technique, combined with fluorescent microscopy, for investigating liposome uptake by endothelial cells.The 90 nm colloidal gold nanoparticles proved to be a noninvasive contrast agent that efficiently improves AFM imaging during the investigation of biological nanoprocesses.

View Article: PubMed Central - PubMed

Affiliation: Division of Cardiology, Department of Internal Medicine, The University of Texas Health Science Center at Houston, 1881 East Road, Houston, TX 77054, USA.

ABSTRACT
Although atomic force microscopy (AFM) has been used extensively to characterize cell membrane structure and cellular processes such as endocytosis and exocytosis, the corrugated surface of the cell membrane hinders the visualization of extracellular entities, such as liposomes, that may interact with the cell. To overcome this barrier, we used 90 nm nanogold particles to label FITC liposomes and monitor their endocytosis on human coronary artery endothelial cells (HCAECs) in vitro. We were able to study the internalization process of gold-coupled liposomes on endothelial cells, by using AFM. We found that the gold-liposomes attached to the HCAEC cell membrane during the first 15-30 min of incubation, liposome cell internalization occurred from 30 to 60 min, and most of the gold-labeled liposomes had invaginated after 2 hr of incubation. Liposomal uptake took place most commonly at the periphery of the nuclear zone. Dynasore monohydrate, an inhibitor of endocytosis, obstructed the internalization of the gold-liposomes. This study showed the versatility of the AFM technique, combined with fluorescent microscopy, for investigating liposome uptake by endothelial cells. The 90 nm colloidal gold nanoparticles proved to be a noninvasive contrast agent that efficiently improves AFM imaging during the investigation of biological nanoprocesses.

No MeSH data available.


Related in: MedlinePlus