Metrological Protocols for Reaching Reliable and SI-Traceable Size Results for Multi-Modal and Complexly Shaped Reference Nanoparticles

Abstract

The study described in this paper was conducted in the framework of the European nPSize project (EMPIR program) with the main objective of proposing new reference certified nanomaterials for the market in order to improve the reliability and traceability of nanoparticle size measurements. For this purpose, bimodal populations as well as complexly shaped nanoparticles (bipyramids, cubes, and rods) were synthesized. An inter-laboratory comparison was organized for comparing the size measurements of the selected nanoparticle samples performed with electron microscopy (TEM, SEM, and TSEM), scanning probe microscopy (AFM), or small-angle X-ray scattering (SAXS). The results demonstrate good consistency of the measured size by the different techniques in cases where special care was taken for sample preparation, instrument calibration, and the clear definition of the measurand. For each characterization method, the calibration process is described and a semi-quantitative table grouping the main error sources is proposed for estimating the uncertainties associated with the measurements. Regarding microscopy-based techniques applied to complexly shaped nanoparticles, data dispersion can be observed when the size measurements are affected by the orientation of the nanoparticles on the substrate. For the most complex materials, hybrid approaches combining several complementary techniques were tested, with the outcome being that the reliability of the size results was improved.

Thymoquinone Nanoparticles (TQ-NPs) in Kidney Toxicity Induced by Ehrlich Ascites Carcinoma (EAC): An In Vivo Study

“…Following centrifugation for about 9000 rcf for 20 minutes at 4°C, the synthesized material was collected and resuspended in 2% sucrose. The shape and size of the particles were confirmed using transmission electron microscope (TEM) (TEM is described in details in the literature 27 , 28 ). The suspension of TQ-PLGA NPs exhibited turbid white color, and the NPs were spherical in shape and 20 nm (10-30 nm) sized with 80% encapsulation efficiency.…”

Strategy for Ensuring the Metrological Traceability of Nanoparticle Size Measurements by SEM

“…Before their sizes can be measured using a Scanning Electron Microscope, the NPs are first deposited onto a silicon substrate (Agar Scientific AGG3390, Stansted, UK) with a spin coater following the protocol described in [ 5 ]. The substrate is functionalized with PLL (Poly-L-Lysine) to enhance the adhesion of the NPs [ 18 , 19 ] (See Figure 1 ).…”

“…The substrate is functionalized with PLL (Poly-L-Lysine) to enhance the adhesion of the NPs [18,19] (See Figure 1). a spin coater following the protocol described in [5]. The substrate is functionalized with PLL (Poly-L-Lysine) to enhance the adhesion of the NPs [18,19] (See Figure 1).…”

“…SEM-based imaging techniques are essentially counting methods and are particularly suitable for determining the size distribution of an NP population. The image acquisition process must be preceded by a specific sample preparation whose objective is to effectively disperse the particles on the substrate in order to limit the agglomerate formation inducing measurement errors [ 5 ]. Then, the isolated particles are measured one by one from a SEM image and the number of particles to be counted to construct a size distribution histogram statistically representative of the population depends on the targeted uncertainty, the polydispersity and the nature of the size distribution (Gaussian, lognormal…) [ 6 ].…”

“…The construction of the latter is essential to detect NP multimodal populations. In this case, each mode is fitted by a Gaussian/lognormal distribution for determining the characteristic parameters [ 5 ]. The most frequently used measurand in SEM measurements is the equivalent circular diameter (ECD), D eq , representing the diameter of a circle that occupies the same two-dimensional surface area as the object to be measured.…”