A Novel Flow-Cytometry-Based Assay for Cellular Uptake Studies of Polyelectrolyte Microcapsules

Small ◽  
2008 ◽  
Vol 4 (10) ◽  
pp. 1763-1768 ◽  
Author(s):  
Maximilian Semmling ◽  
Oliver Kreft ◽  
Almudena Muñoz Javier ◽  
Gleb B. Sukhorukov ◽  
Josef Käs ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cellular Uptake ◽  
Polyelectrolyte Microcapsules ◽  
Uptake Studies

scholarly journals In Vitro Cellular Uptake Studies of Self-Assembled Fluorinated Nanoparticles Labelled with Antibodies

Nanomaterials ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1906
Author(s):  
Mona Atabakhshi-Kashi ◽  
Mónica Carril ◽  
Hossein Mahdavi ◽  
Wolfgang J. Parak ◽  
Carolina Carrillo-Carrion ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cellular Uptake ◽  
Hydrophobic Interactions ◽  
Intrinsic Properties ◽  
Wide Range ◽  
Self Assembled ◽  
Uptake Studies ◽  
Targeting Ability ◽  
Fluorinated Nanoparticles

Nanoparticles (NPs) functionalized with antibodies (Abs) on their surface are used in a wide range of bioapplications. Whereas the attachment of antibodies to single NPs to trigger the internalization in cells via receptor-mediated endocytosis has been widely studied, the conjugation of antibodies to larger NP assemblies has been much less explored. Taking into account that NP assemblies may be advantageous for some specific applications, the possibility of incorporating targeting ligands is quite important. Herein, we performed the effective conjugation of antibodies onto a fluorescent NP assembly, which consisted of fluorinated Quantum Dots (QD) self-assembled through fluorine–fluorine hydrophobic interactions. Cellular uptake studies by confocal microscopy and flow cytometry revealed that the NP assembly underwent the same uptake procedure as individual NPs; that is, the antibodies retained their targeting ability once attached to the nanoassembly, and the NP assembly preserved its intrinsic properties (i.e., fluorescence in the case of QD nanoassembly).

scholarly journals EFFECTS OF POLYMERIC NANOPARTICLE SURFACE PROPERTIES ON INTERACTION WITH BRAIN TUMOR ENVIRONMENT

Nano LIFE ◽  
2013 ◽  
Vol 03 (04) ◽  
pp. 1343003 ◽  
Author(s):  
BRANDON MATTIX ◽  
THOMAS MOORE ◽  
OLGA UVAROV ◽  
SAMUEL POLLARD ◽  
LAUREN O'DONNELL ◽  
...  
Keyword(s):  
Human Brain ◽  
Surface Charge ◽  
Cellular Uptake ◽  
Drug Clearance ◽  
Cellular Interaction ◽  
Normal Tissues ◽  
Poly Ethylene ◽  
Uptake Studies ◽  
Effect Of Surface

Current chemotherapy treatments are limited by poor drug solubility, rapid drug clearance and systemic side effects. Additionally, drug penetration into solid tumors is limited by physical diffusion barriers [e.g., extracellular matrix (ECM)]. Nanoparticle (NP) blood circulation half-life, biodistribution and ability to cross extracellular and cellular barriers will be dictated by NP composition, size, shape and surface functionality. Here, we investigated the effect of surface charge of poly(lactide)-poly(ethylene glycol) NPs on mediating cellular interaction. Polymeric NPs of equal sizes were used that had two different surface functionalities: negatively charged carboxyl ( COOH ) and neutral charged methoxy ( OCH 3). Cellular uptake studies showed significantly higher uptake in human brain cancer cells compared to noncancerous human brain cells, and negatively charged COOH NPs were uptaken more than neutral OCH 3 NPs in 2D culture. NPs were also able to load and control the release of paclitaxel (PTX) over 19 days. Toxicity studies in U-87 glioblastoma cells showed that PTX-loaded NPs were effective drug delivery vehicles. Effect of surface charge on NP interaction with the ECM was investigated using collagen in a 3D cellular uptake model, as collagen content varies with the type of cancer and the stage of the disease compared to normal tissues. Results demonstrated that NPs can effectively diffuse across an ECM barrier and into cells, but NP mobility is dictated by surface charge. In vivo biodistribution of OCH 3 NPs in intracranial tumor xenografts showed that NPs more easily accumulated in tumors with less collagen. These results indicate that a robust understanding of NP interaction with various tumor environments can lead to more effective patient-tailored therapies.

scholarly journals Epigallocatechin Gallate-Gold Nanoparticles Exhibit Superior Antitumor Activity Compared to Conventional Gold Nanoparticles: Potential Synergistic Interactions

Nanomaterials ◽  
2019 ◽  
Vol 9 (3) ◽  
pp. 396 ◽  
Author(s):  
Suhash Chavva ◽  
Sachin Deshmukh ◽  
Rajashekhar Kanchanapally ◽  
Nikhil Tyagi ◽  
Jason Coym ◽  
...  
Keyword(s):  
Gold Nanoparticles ◽  
Antitumor Activity ◽  
Cancer Cells ◽  
Cellular Uptake ◽  
Nuclear Translocation ◽  
Epigallocatechin Gallate ◽  
Drug Carriers ◽  
Water Soluble ◽  
Tetrazolium Salt ◽  
Uptake Studies

Epigallocatechin gallate (EGCG) possesses significant antitumor activity and binds to laminin receptors, overexpressed on cancer cells, with high affinity. Gold nanoparticles (GNPs) serve as excellent drug carriers and protect the conjugated drug from enzymatic metabolization. Citrate-gold nanoparticles (C-GNPs) and EGCG-gold nanoparticles (E-GNPs) were synthesized by reduction methods and characterized with UV-visible spectroscopy, transmission electron microscopy (TEM), and dynamic light scattering (DLS). Cytotoxicity of citrate, EGCG, C-GNPs, and E-GNPs was evaluated by the water-soluble tetrazolium salt (WST-1) assay. Nanoparticle cellular uptake studies were performed by TEM and atomic absorption spectroscopy (AAS). Dialysis method was employed to assess drug release. Cell viability studies showed greater growth inhibition by E-GNPs compared to EGCG or C-GNPs. Cellular uptake studies revealed that, unlike C-GNPs, E-GNPs were taken up more efficiently by cancerous cells than noncancerous cells. We found that E-GNP nanoformulation releases EGCG in a sustained fashion. Furthermore, data showed that E-GNPs induced more apoptosis in cancer cells compared to EGCG and C-GNPs. From the mechanistic standpoint, we observed that E-GNPs inhibited the nuclear translocation and transcriptional activity of nuclear factor-kappaB (NF-κB) with greater potency than EGCG, whereas C-GNPs were only minimally effective. Altogether, our data suggest that E-GNPs can serve as potent tumor-selective chemotoxic agents.

Biological activities and cellular uptake studies of fluorescent derivatives of indole alkaloid tumor promoter teleocidin

1989 ◽  
Vol 43 (3) ◽  
pp. 513-519 ◽  
Author(s):  
Kazuhiro Irie ◽  
Shigenori Okuno ◽  
Koichi Koshimizu ◽  
Harukuni Tokuda ◽  
Hoyoku Nishino ◽  
...  
Keyword(s):  
Cellular Uptake ◽  
Biological Activities ◽  
Indole Alkaloid ◽  
Tumor Promoter ◽  
Uptake Studies ◽  
Derivatives Of ◽  
Fluorescent Derivatives

scholarly journals Understanding the Underlying Immune Response in HIT: Uptake of PF4/Heparin Complexes By Monocytes & Dendritic Cells

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4197-4197
Author(s):  
Manali V. Joglekar ◽  
Sanjay Khandelwal ◽  
Mortimer Poncz ◽  
Lubica Rauova ◽  
Gowthami M Arepally
Keyword(s):  
Flow Cytometry ◽  
Immune Response ◽  
Dendritic Cells ◽  
Cellular Uptake ◽  
Dependent Manner ◽  
Cellular Interactions ◽  
Platelet Factor ◽  
Cellular Activation ◽  
Late Endosomes ◽  
Positively Charged

Abstract Platelet Factor 4 (PF4), a strongly positive-charged small protein, and the negatively-charged polymer heparin (Hep) form ultra-large complexes (ULCs) that have several unusual features including their remarkable stability and their ability to elicit a robust antibody response in vivo (Rauova L; Blood 2005 and Suvarna S, Blood 2005). Recently, we and others have shown that similar complexes of another positively charged protein, protamine sulfate with heparin can also lead to a clinically relevant immune response. To better understand the cellular basis for PF4/Hep antibody formation, we investigated mechanisms of cellular interactions and uptake of PF4 and PF4/Hep ULCs. For these studies, we examined cellular uptake of unlabeled or labeled PF4, heparin, or PF4/Hep using monocytes (PBMCs), dendritic cells (DCs) and/or neutrophils derived from peripheral blood. For these studies, cells were incubated with varying concentrations of unlabeled or fluorescently-labeled antigen (PF4, Hep or PF4/Hep ULCs). In cellular studies using unlabeled antigen, uptake was detected by fluorescently-labeled KKO, a monoclonal antibody to PF4/Hep complexes. Cellular uptake was visualized by confocal microscopy or flow cytometry. In initial studies, we defined the time course of uptake. As shown in Figure 1, we demonstrate that PF4/Hep-FITC ULCs are taken up by PBMCs in a time-dependent manner, with maximal uptake occurring between 12-24 hours (Figure 1, only 24 hour time point shown). This uptake is independent of the fluorescent label, as labeled or unlabeled intracellular PF4/Hep ULCs were readily visualized by KKO-AF647. To examine the effect of Hep on PF4 uptake, PBMCs were incubated with unlabeled PF4 alone or in the presence of increasing concentrations of Hep-FITC (0.1-2.5 U/mL). As shown in Figure 2A, Hep markedly enhances the efficiency of cellular uptake of PF4 in a Hep-dependent manner. Increased number of intracellular vesicles containing labeled PF4/Hep-FITC was noted at Hep-FITC concentration of 0.25-1 U/mL (Figure 2B; fluorescent vesicles/cell: 0.6 ± 0.22 for 0.35 U/mL and 0.5 ± 0.26 for 1U/mL) as compared to PF4 alone (25 µg/mL; number of fluorescent vesicles/cell: 0.35 ± 0.07). On examining the uptake of ULCs by other phagocytic cells, we could not demonstrate PF4/Hep uptake by neutrophils, suggesting that only monocytes/DCs provide clearance of complexes. Cellular uptake of Hep-containing ULCs was not limited to complexes of PF4 and heparin but also other positively-charged proteins, as intracellular complexes could be demonstrated when Hep-FITC was incubated with murine PF4, protamine, or lysozyme to form corresponding protein/Hep-FITC ULCs. This uptake was an active process of monocytes as PF4/Hep ULC endocytosis was inhibited by 4C, and cytochalasin D, an actin polymerization inhibitor and was associated with cellular activation and expression of MHC II and CD83 co-stimulatory molecules as shown by flow cytometry (Figure 3). Finally, co-staining with KKO and lysosomal associated membrane protein-2 (LAMP-2) localized intact PF4/Hep ULCs into late endosomes. Taken together, these studies demonstrate that PF4/Hep and other protein/heparin ULCs are taken up actively by monocytes and/or DCs, intact ULCs can be detected in late endosomes and uptake is accompanied by cellular activation. These studies establish a distinct role for heparin in increasing the uptake and cellular activation of PF4 and other positively charged complexes. These studies additionally provide insights into why the majority of clinical cases of HIT occur in the wake of heparin exposure. Disclosures Arepally: TEVA Pharma: Consultancy.

Synthesis of Water-Soluble Thioglycosylated trans-A2B2 Type Porphyrins: Cellular Uptake Studies and Photodynamic Efficiency

2020 ◽  
Vol 85 (10) ◽  
pp. 6309-6322 ◽  
Author(s):  
Vijayalakshmi Pandey ◽  
Md Kausar Raza ◽  
Pooja Joshi ◽  
Iti Gupta
Keyword(s):  
Cellular Uptake ◽  
Water Soluble ◽  
Uptake Studies
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