Timing, hosts and locations of (grouped) events of NanoImpactNet

NanoSafetyCluster - Compendium 2012
Figure 2: Reference material: Tomato soup spiked with silica
Experiments to stabilise meat, the target matrix for the silver
particles, have been executed: freezing is not possible because of
potential change and agglomeration of particles. Steam
sterilisation was also not successful, but gamma-irradiation was
found to prevent degradation. Ag-nanoparticles in aqueous
suspension remained unchanged by irradiation, hinting that it
should not cause any problems also for the matrix material.
Currently, meat spiked with Ag nanoparticles, mimicking e.g.
diffusion from Ag-impregnated cutting boards, is being prepared.
Labelled particles have been synthesised and characterised: Silica
nanoparticles containing a small amount of GeO2 and Ag
nanoparticles containing a small amount of Au were produced. The
materials were characterised with respect to their particle size,
dispersion behaviour and Au/Ge mass fraction. The particles show
sufficient stability and are intended as internal standards and/or as
recovery control for the methods being developed
Finally, a common validation approach for the determination of
nanoparticles in food was developed. The proposed approach will
be published as a basis for discussion among scientific and
regulatory communities and the iterative development and
advancement of robust and widely accepted validation guidelines.
4.2 WP2: Rapid imaging and screening methods
OBJECTIVES
Presumably, many foods will not contain any engineered
nanoparticles. Applying rapid, cost-efficient and robust methods to
distinguish the samples which actually contain engineered
nanoparticles from the majority which doesn’t, would allow to
focus more laborious quantitative methods on those samples. The
objective of WP2 is to develop such rapid analytical methods,
based on imaging and screening techniques, for providing
qualitative and semi-quantitative data on engineered nanoparticles
in different food matrices. The developed methods should enable a
rapid decision if any target particles are present or absent in a food
sample.
• Sample preparation methodology tailored to imaging and
screening methods for engineered nanoparticles in foods
• A simplified electron microscopic imaging tool with
automated smart image analysis for the rapid detection
of engineered nanoparticles presence in different food
matrices
• Screening assays for engineered nanoparticles in sensor
and ELISA format
ACTIVITIES
In order that the developed techniques can be applied broadly in
the future, the work package will explore a range of engineered
nanoparticle types that are relevant to food such as: metal-based
(Ag), metal oxides (SiO2), and organic nano-carrier systems.
Electron microscopy: A limited number of imaging methodologies
(SEM, TEM, including inherent characterisation of elemental
composition by EDX, EELS) will be compared initially for aqueous
engineered nanoparticles dispersions. The most suited will be
selected for further method development. A major challenge will
be the preparation of the food materials for analysis. The work
package will therefore explore sample preparation techniques for
a range of matrices starting with non-complex systems (i.e. water),
and finally moving to more complex matrices. Work will be focused
on easy to use and low-cost techniques e.g. resin embedding. For
more complex samples a range of more sophisticated techniques
will be available, e.g. capsules to enable imaging under fully liquid
conditions. Finally the most successful sample preparation and
detection methods for different engineered nanoparticle types will
be combined into fully validated methods. In order to achieve
automation and high throughput automated object-based image
analysis will be explored and further developed.
Screening assays: Two approaches will be followed:
(i) an ELISA approach for engineered nanoparticles for direct
implementation in basic food labs,
(ii) a sensor approach based either on bio- or physico-chemical
recognition of functionalised, encapsulate or metal(oxide)
engineered nanoparticles, respectively, for automated high
throughput analysis. Both methods will be validated according to
the standards for screening methods.
The validation will include the analysis of a limited number of real
samples from the market, claiming or suspect to contain
engineered nanoparticles.
RESULTS
Concerning electron microscopy, a number of sample preparation
methods for nanoparticle samples both in liquid dispersions and
food matrices have been tested. These include air drying, blotting,
freeze drying, ultracentrifugation, chemical drying, resin
embedding, and freezing for imaging in liquid conditions. Of the
methods tested so far, ultracentrifugation has been identified as
the most appropriate method for preparation of samples at low
concentrations of ENPs. The method minimises aggregation of
ENPs during processing. Blotting is another rapid and cheap
method for working with untreated samples as it does not cause
any major changes to ENPs in comparison with other drying-ongrid
methods.
An initial prototype object-based software system for analysis of
nanoparticle EM images in food matrices has been developed for
semi-automated analysis of electron microscopy images of
nanoparticles in food matrices.
Antibodies have been raised against cross-linked gelatine
nanoparticles that are a component of nano-sized carriers for
136 Compendium of Projects in the European NanoSafety Cluster