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Quantification of Al₂O₃ nanoparticles in human cell lines applying inductively coupled plasma mass spectrometry (neb-ICP-MS, LA-ICP-MS) and flow cytometry-based methods.

ABSTRACT: In order to quantify and compare the uptake of aluminum oxide nanoparticles of three different sizes into two human cell lines (skin keratinocytes (HaCaT) and lung epithelial cells (A549)), three analytical methods were applied: digestion followed by nebulization inductively coupled plasma mass spectrometry (neb-ICP-MS), direct laser ablation ICP-MS (LA-ICP-MS), and flow cytometry. Light and electron microscopy revealed an accumulation and agglomeration of all particle types within the cell cytoplasm, whereas no particles were detected in the cell nuclei. The internalized Al₂O₃ particles exerted no toxicity in the two cell lines after 24 h of exposure. The smallest particles with a primary particle size (x_BET) of 14 nm (Alu1) showed the lowest sedimentation velocity within the cell culture media, but were calculated to have settled completely after 20 h. Alu2 (x_BET = 111 nm) and Alu3 (x_BET = 750 nm) were calculated to reach the cell surface after 7 h and 3 min, respectively. The internal concentrations determined with the different methods lay in a comparable range of 2-8 µg Al₂O₃/cm² cell layer, indicating the suitability of all methods to quantify the nanoparticle uptake. Nevertheless, particle size limitations of analytical methods using optical devices were demonstrated for LA-ICP-MS and flow cytometry. Furthermore, the consideration and comparison of particle properties as parameters for particle internalization revealed the particle size and the exposure concentration as determining factors for particle uptake.

SUBMITTER: Bohme S

PROVIDER: S-EPMC4176630 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The Centers for Disease Control and Prevention developed a biomonitoring method to rapidly and accurately quantify chromium and cobalt in human whole blood by ICP-MS. Many metal-on-metal hip implants which contain significant amounts of chromium and cobalt are susceptible to metal degradation. This method is used to gather population data about chromium and cobalt exposure of the U.S. population that does not include people that have metal-on-metal hip implants so that reference value can be established for a baseline level in blood. We evaluated parameters such as; helium gas flow rate, choice and composition of the diluent solution for sample preparation, and sample rinse time to determine the optimal conditions for analysis. The limits of detection for chromium and cobalt in blood were determined to be 0.41 and 0.06 ?g/L, respectively. Method precision, accuracy, and recovery for this method were determined using quality control material created in-house and historical proficiency testing samples. We conducted experiments to determine if quantitative changes in the method parameters affect the results obtained by changing four parameters while analyzing human whole blood spiked with National Institute of Standard and Technology traceable materials: the dilution factor used during sample preparation, sample rinse time, diluent composition, and kinetic energy discrimination gas flow rate. The results at the increased and decreased levels for each parameter were statistically compared to the results obtained at the optimized parameters. We assessed the degree of reproducibility obtained under a variety of conditions and evaluated the method's robustness by analyzing the same set of proficiency testing samples by different analysts, on different instruments, with different reagents, and on different days. The short-term stability of chromium and cobalt in human blood samples stored at room temperature was monitored over a time period of 64 hours by diluting and analyzing samples at different time intervals. The stability of chromium and cobalt post-dilution was also evaluated over a period of 48 hours and at two storage temperatures (room temperature and refrigerated at 4°C). The results obtained during the stability studies showed that chromium and cobalt are stable in human blood for a period of 64 hours.

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Quantification of Al₂O₃ nanoparticles in human cell lines applying inductively coupled plasma mass spectrometry (neb-ICP-MS, LA-ICP-MS) and flow cytometry-based methods.

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Quantification of Al₂O₃ nanoparticles in human cell lines applying inductively coupled plasma mass spectrometry (neb-ICP-MS, LA-ICP-MS) and flow cytometry-based methods.