ABSTRACT
Antibodies are a class of molecules widely used in bioengineering and nanomedicine for applications involving protein recognition and targeting. Here we report an efficient method for universal conjugation of antibodies to lipid-coated nanoparticles using radially oriented FcγRIs. This method is performed in physiological solution with no additional coupling reagents, thereby avoiding problems with antibody stability and functionality. Coupling to the Fc region of the antibody avoids aggregation and polymerization allowing high yield. In addition, the antibody is oriented perpendicular to the surface so that the binding sites are fully functional. Using this method we demonstrate quantitative profiling of a panel of four membrane-bound cancer biomarkers (claudin-4, mesothelin, mucin-4, and cadherin-11) on four cell lines (Panc-1, MIA PaCa-2, Capan-1, and HPDE). We show that by designing the lipid coating to minimize aggregation and nonspecific binding, we can obtain absolute values of biomarker expression levels as number per unit area on the cell surface. This method is applicable to a wide range of technologies, including solution based protein detection assays and active targeting of cell surface membrane biomarkers.
Subject(s)
Immunoconjugates/chemistry , Nanoparticles/chemistry , Pancreatic Neoplasms/diagnosis , Receptors, IgG/chemistry , Animals , Biomarkers, Tumor/analysis , Cadherins/analysis , Cell Line, Tumor , Claudin-4/analysis , GPI-Linked Proteins/analysis , Humans , Mesothelin , Mice , Mucin-4/analysis , Recombinant Proteins/chemistryABSTRACT
The respiratory system is one of the portals of entry into the body, and hence inhalation of engineered nanomaterials is an important route of exposure. The broad range of physicochemical properties that influence biological responses necessitate the systematic study to contribute to understanding occupational exposure. Here, we report on the influence of nanoparticle charge and dose on human airway epithelial cells, and show that this platform can be used to evaluate consequences of exposure to engineered nanomaterials.
Subject(s)
Bronchi/cytology , Epithelial Cells/metabolism , Quantum Dots/chemistry , Quantum Dots/metabolism , Biological Transport , Cadmium Compounds/chemistry , Cells, Cultured , Epithelial Cells/drug effects , Humans , Selenium Compounds/chemistry , Surface PropertiesABSTRACT
Little is known about engineered nanoparticles (NPs) exposures on oysters. As sessile filter feeders, oysters are likely to be exposed to NPs suspended in the water column with unknown effects of NP exposure on oyster functioning. Our study indicates that waterborne NPs alter oyster hemocyte phagocytosis dynamics, an indication of sub-lethal effects of NP exposures. Silver NPs, titanium dioxide (TiO(2)) NPs, and silver nitrate exposures reduced phagocytosis compared to the control. Increasing TiO(2) NPs and silver nitrate concentrations reduced phagocytosis. Silver NPs, up to 120ppb, increased phagocytosis, but higher concentrations reduced phagocytosis.
Subject(s)
Hemocytes/drug effects , Metal Nanoparticles/toxicity , Phagocytosis/drug effects , Water Pollutants, Chemical/toxicity , Animals , Crassostrea , Dose-Response Relationship, Drug , Pilot Projects , Silver/toxicityABSTRACT
Applications in nanomedicine, such as diagnostics and targeted therapeutics, rely on the detection and targeting of membrane biomarkers. In this article we demonstrate absolute quantitative profiling, spatial mapping, and multiplexing of cancer biomarkers using functionalized quantum dots (QDs). We demonstrate highly selective targeting molecular markers for pancreatic cancer with extremely low levels of nonspecific binding. We confirm that we have saturated all biomarkers on the cell surface, and, in conjunction with control experiments, extract absolute quantitative values for the biomarker density in terms of the number of molecules per square micron on the cell surface. We show that we can obtain quantitative spatial information of biomarker distribution on a single cell, important because tumors' cell populations are inherently heterogeneous. We validate our quantitative measurements (number of molecules per square micron) using flow cytometry and demonstrate multiplexed quantitative profiling using color-coded QDs. FROM THE CLINICAL EDITOR: This paper demonstrates a nice example for quantum dot-based molecular targeting of pancreatic cancer cells for advanced high sensitivity diagnostics and potential future selective therapeutic purposes.
Subject(s)
Antibodies , Immunoconjugates , Nanomedicine/methods , Pancreas/pathology , Pancreatic Neoplasms/diagnosis , Quantum Dots , Biomarkers, Tumor/analysis , Cell Line, Tumor , Humans , Lipids/chemistry , Pancreatic Neoplasms/pathology , Pancreatic Neoplasms/therapyABSTRACT
The water solubilization of nanoparticles is key for many applications in biomedicine. Despite the importance of surface functionalization, progress has been largely empirical and very few systematic studies have been performed. Here we report on the water solubilization of quantum dots using lipid encapsulation. We systematically evaluate the monodispersity, zeta potential, stability, and quantum yield for quantum dots encapsulated with single and double acyl-chain lipids, pegylated double acyl-chain lipids, and single alkyl-chain surfactant molecules with charged head groups. We show that charged surfactants and pegylated lipids are important to obtain monodisperse suspensions with high yield and excellent long-term stability. FROM THE CLINICAL EDITOR: This study reports on solubilization of nanoparticles in water, a key, but often neglected aspect for biomedical applications. The authors demonstrate that charged surfactants and PEGylated lipids are important to obtain monodisperse suspensions with high yield and long-term stability.
