Production and characterization of antibodies against crosslinked gelatin nanoparticles and first steps toward developing an ELISA screening kit

Nanotechnologies are finding a growing range of applications in the food sector. Nanoparticles are used notably to add vitamins and other nutrients to foods and beverages without affecting taste and color. They are also used to develop new tastes, preserve food texture, control the release of flavors, improve the bioavailability of compounds such as antioxidants and vitamins, and monitor freshness with nanosensors. Crosslinked gelatin nanoparticles are a component of nano-sized carriers for nutrient and supplement delivery in foods and related products. This paper describes the production and characterization of polyclonal antibodies against gelatin nanoparticles. Two immunization schemes were investigated: subcutaneous injection with and without a first intravenous injection. Two enzyme-linked immunosorbent assay formats were used to characterize the antibodies: an inhibition format with an antigen-coated plate for detection of the immune response and a sandwich format for development of the method. The antibodies showed good sensitivity with an IC50 equal to 0.11 ng mL-1 using indirect ELISA format and a good specificity for the nanomaterials, without significant cross-reactivity against native gelatin. The limit of detection was determined—0.42, 0.27, 0.26, and 0.24 µg mL-1 for apple, orange juice, milk, and soft drink matrices, respectively. ELISA technology offers rapid, low-cost assays for screening foods, feeds, and beverages. We have studied a prototype ELISA for detection of gelatin-based nanocarrier systems. Fruit juices, milk, and a soft drink were the matrices selected for assay development

Results of an interlaboratory method performance study for the size determination and quantification of silver nanoparticles in chicken meat by single-particle-inductively coupled plasma-mass spectrometry (sp-ICP-MS)

Single-particle inductively coupled plasma mass spectrometry (sp-ICP-MS) promises fast and selective determination of nanoparticle size and number concentrations. While several studies on practical applications have been published, data on formal, especially interlaboratory validation of sp-ICP-MS is sparse. An international interlaboratory study was organized to determine repeatability and reproducibility of the determination of the median particle size and particle number concentration of Ag nanoparticles (AgNPs) in chicken meat, Ten laboratories from the European Union, USA and Canada, determined particle size and particle number concentration of two chicken meat homogenates spiked with polyvinylpyrrolidone (PVP) stabilized AgNPs. For the determination of the median particle diameter, repeatability standard deviations of 2 % and 5 % were determined, and reproducibility standard deviations were 15 % and 25 %, respectively. Determination of the particle number concentration was significantly less precise, with repeatability standard deviations of 7 % and 18 % and reproducibility standard deviations of 70 and 80 %.JRC.F.6-Reference Material

Production and characterization of antibodies against crosslinked gelatin nanoparticles and first steps toward developing an ELISA screening kit

Nanotechnologies are finding a growing range of applications in the food sector. Nanoparticles (NPs) are used notably to add vitamins and other nutrients to foods and beverages without affecting taste and color. They are also used to develop new tastes, preserve food texture, control the release of flavors, improve the bioavailability of compounds such as antioxidants and vitamins, and monitor freshness with nanosensors. Crosslinked gelatin nanoparticles are a component of nano-sized carriers for nutrient and supplement delivery in foods and related products. This paper describes the production and characterization of polyclonal antibodies against gelatin nanoparticles. Two immunization schemes were investigated: subcutaneous injection with and without a first intravenous injection. Two enzyme-linked immunosorbent assay formats were used to characterize the antibodies: an inhibition format with an antigen-coated plate for detection of the immune response and a sandwich format for development of the method. The antibodies showed good specificity for the nanomaterials and good sensitivity, without significant cross-reactivity against native gelatin. ELISA technology offers rapid, low-cost assays for screening foods, feeds, and beverages. We have studied a prototype ELISA for detection of gelatin-based nanocarrier systems. Fruit juices, milk, and a soft drink were the matrices selected for assay development.JRC.D.2-Standards for Innovation and sustainable Developmen

Validation of methods for the detection and quantification of engineered nanoparticles in food

The potential impact of nanomaterials on the environment and on human health has already triggered legislation requiring labelling of products containing nanoparticles. However, so far, no validated analytical methods for the implementation of this legislation exist. This paper outlines a generic approach for the validation of methods for detection and quantification of nanoparticles in food samples. It proposes validation of identity, selectivity, precision, working range, limit of detection and robustness, bearing in mind that each “result” must include information about the chemical identity, particle size and mass or particle number concentration. This has an impact on testing for selectivity and trueness, which also must take these aspects into consideration. Selectivity must not only be tested against matrix constituents and other nanoparticles, but it shall also be tested whether the methods apply equally well to particles of different suppliers. In trueness testing, information whether the particle size distribution has changed during analysis is required. Results are largely expected to follow normal distributions due to the expected high number of particles. An approach of estimating measurement uncertainties from the validation data is given

Feasibility of the development of reference materials for the detection of Ag nanoparticles in food: neat dispersions and spiked chicken meat

The feasibility of producing colloidal silver nanoparticle reference materials and silver nanoparticle spiked reference matrix materials was investigated. Two concentrations of PVP-coated silver nanoparticle dispersions were evaluated and used to spike chicken meat, with the aim of producing a set of reference materials to support the development of analytical methods for the detection and quantification of nanoparticles in food. Aqueous silver nanoparticle (AgNP) dispersions were evaluated for their homogeneity of mass fraction and particle size and found sufficiently homogeneous to be used as reference materials.Stability studies at 4 C, 18 C and 60 C demonstrated sufficient short- and long-term stability, although particle size decreases in a linear fashion at 60 C. The AgNP dispersions were characterized for total Ag mass fraction by ICP-OES, dissolved Ag content by ultrafiltration-ICPMS, as well as AgNP particle size by dynamic light scattering, transmission electron microscopy (TEM) and gasphase electrophoretic molecular mobility analysis. Chicken breasts were homogenized by cryo-milling and spiked with aqueous AgNP dispersions. Rapid freezing over liquid nitrogen resulted in homogeneous and stable materials. The spiked chicken materials were characterized for their total Ag mass fraction by neutron activation analysis and for the AgNP particle size by TEM and single-particle inductively coupled plasma mass spectrometry. The observed differences in particle sizes between the spiked chicken samples and the original silver dispersions indicate relevant matrix effects. The materials demonstrate that production and characterization of reference materials for the detection and quantification of silver nanoparticles in meat are feasible, but challenges especially in assessing stability and having sufficiently precise methods for assessment of homogeneity and stability remain.JRC.D.2-Standards for Innovation and sustainable Developmen

Feasibility of the development of reference materials for the detection of Ag nanoparticles in food: neat dispersions and spiked chicken meat

The feasibility of producing colloidal silver nanoparticle reference materials and silver nanoparticle spiked reference matrix materials was investigated. Two concentrations of PVP-coated silver nanoparticle dispersions were evaluated and used to spike chicken meat, with the aim of producing a set of reference materials to support the development of analytical methods for the detection and quantification of nanoparticles in food. Aqueous silver nanoparticle (AgNP) dispersions were evaluated for their homogeneity of mass fraction and particle size and found sufficiently homogeneous to be used as reference materials. Stability studies at 4 °C, 18 °C and 60 °C demonstrated sufficient short- and long-term stability, although particle size decreases in a linear fashion at 60 °C. The AgNP dispersions were characterized for total Ag mass fraction by ICP-OES, dissolved Ag content by ultrafiltration-ICP-MS, as well as AgNP particle size by dynamic light scattering, transmission electron microscopy (TEM) and gas-phase electrophoretic molecular mobility analysis. Chicken breasts were homogenized by cryo-milling and spiked with aqueous AgNP dispersions. Rapid freezing over liquid nitrogen resulted in homogeneous and stable materials. The spiked chicken materials were characterized for their total Ag mass fraction by neutron activation analysis and for the AgNP particle size by TEM and single-particle inductively coupled plasma mass spectrometry. The observed differences in particle sizes between the spiked chicken samples and the original silver dispersions indicate relevant matrix effects. The materials demonstrate that production and characterization of reference materials for the detection and quantification of silver nanoparticles in meat are feasible, but challenges especially in assessing stability and having sufficiently precise methods for assessment of homogeneity and stability remain