Icap qc icp ms

Metal analysis was performed at the Northwestern University Quantitative Bio-element Imaging Center generously supported by NASA Ames Research Center NNA06CB93G. Total cell pellet and extracellular copper were assessed by inductively-coupled plasma mass spectrometry (Thermo Fisher Scientific iCap Qc ICP-MS). Cells were seeded at 300,000 cells per well and stimulated for 1, 6, 12, and 24 h. Media were collected and spun at 500 × g for 10 min. A total of 250 µL media were added to pre-weighed metal-free 15-ml conical tubes (89049-170, VWR), and 250 µl analytical grade 70% nitric acid (A509P500, Thermo Fisher Scientific) was added to the mixture and left at room temperature for 24 h. Cells were washed two times with cold PBS and digested in 250 µl analytical grade 70% nitric acid. After 24 h of acid digestion, 225 µl of cell lysate was transferred to pre-weighed metal-free 15-ml conical tubes. All samples were diluted to 5 ml with 3% analytical grade nitric acid. Copper and phosphorus were assessed by ICP-MS. Intracellular copper concentration is expressed as the ratio of concentrations of copper over phosphorus, while copper concentration in media samples is expressed as the concentration of copper over the mass of the sample.

ICP-MS measurements were performed using Thermo Scientific iCAP Qc ICP-MS (Thermo Scientific, Bremen, Germany) spectrometer with operational software Qtegra. For Ru determination the instrument was adjusted for optimum performance in He KED (kinetic energy discrimination) mode using the supplied autotune protocols. For Os determination the instrument was adjusted for optimum performance standard no gas mode using the supplied autotune protocols. The instrumental operating conditions for ICP-MS are shown in Table 2.
Analytical blanks were run in the same way as the samples, and concentrations were determined using standard solutions prepared in the same acid matrix. The standard for the instrument calibration was prepared on the basis of ruthenium, plasma standard solution, Specpure®, Ru 1000 μg mL⁻¹ and osmium, plasma standard solution, Specpure, Os 1000 μg mL⁻¹ certified reference solutions ICP standard purchased from Alfa Aesar GmbH & Co KG (Germany).

PBS perfused mouse brains were dissected and immediately embedded in Optimal Cutting Temperature (OCT) mounting media (Tissue Tek) in cryomolds (Tissue Tek), then flash-frozen in a liquid nitrogen/isopentane bath and stored at -80° C until sectioning. The embedded brains were sectioned into 20 μm slices using a (Leica Cryostat CM 3050S). The slices were air-dried and stored at room temperature until analysis. Laser ablation was performed on an NWR213 laser with a TV2 sample chamber (ESI, Bozeman, MT) using the following parameters: Spot size: 6 μm; Fluence: 3.1 J cm-2; Stage speed: 15 μm s-1; Firing rate: 20 Hz; He flow: 800 mL min-1; Pattern spacing: 6 μm. The ablated material was introduced by Helium flow into an iCAP-Qc ICP-MS (Thermo Fisher) and analyzed for 63Cu using a 0.4 sec dwell time in standard acquisition mode. The resulting mass spectrometry traces and laser log files were processed in Igor Pro using the Iolite application with a custom matrix.

The morphologies of UCPP and UCNPs were examined by transmission electronic microscopy (TEM, Tecnai G2spirit Biotwin) at an accelerating voltage of 120 kV. The platinum content obtained outside of the dialysis bags in drug release experiments was measured on Inductively Coupled Plasma Mass Spectrometer (ICP-MS, iCAP Qc-ICPMS, Thermoscientific, USA) and Inductively Coupled Plasma Optical Emission Spectrometer (ICP-OES, iCAP 6300, Thermoscientific, USA). ¹H NMR spectra were measured by a Unity-300 MHz NMR spectrometer (Bruker) at room temperature. UV-vis absorption spectra were taken on a Milton Ray Spectronic 3000 array spectrophotometer. Photoluminescence (PL) spectra were measured on a Perkin-Elmer LS-55 spectro-fluorometer. Mass Spectroscopy (ESI-MS) measurements were performed on a Quattro Premier XE system (Waters) equipped with an electrospray interface (ESI). Size and size distribution of micelles were determined by DLS (Zetasizer nano ZS, Malvern, UK). Fourier transform infrared (FTIR) spectra were recorded on a Paragon 1,000 instrument by KBr sample holder method. Oxygen concentration was measured by a oxygen dissolving meter (HI-2400, HANNA, Italy).

The tin and platinum uptake analysis was performed on MCF-7 cells with the aid of inductively coupled plasma mass spectrometry (ICP-MS). The cells were seeded in T25 flasks and allowed to grow. After 24 h, the exponentially growing cells were treated with 10 mL of Ph₃SnL1 and cisplatin in concentrations corresponding to their IC₅₀ values for 24 h, and thereafter trypsinised and collected by centrifugation at 1200 rpm for 8 min. Using 5 mL of ice-cold PBS the cells were washed and the metal uptake concentration in prepared samples was determined according to the previously described procedure [40 (link)]. A Thermo Scientific iCAP Qc ICP-MS (Thermo Scientific, Bremen, Germany) instrument using the supplied autotune protocols and operational software Qtegra was employed for metal uptake determination. The measurements were performed on isotopes ¹¹⁹Sn, ¹²⁰Sn, ¹⁹⁵Pt, and ¹⁹⁸Pt and the concentrations of metals were expressed as ppb (µg/L).

Sample preparation for the measurement of intracellular Ru/Os accumulation using ICP-MS: Ru/Os accumulation was analyzed in A549 cells with ICP-MS using Thermo Scientific iCAP Qc ICP-MS (Thermo Scientific, Bremen, Germany).50 (link) A549 cells were seeded into a 25 cm² dish (Thermo Scientific Nunc™) and treated with the complexes 4 and 8 at concentrations equal to 0.5×IC₅₀. After 6 and 24 h, cells were harvested by scraping, washed with ice-cold PBS and collected by centrifugation at 778 g for 10 min.
Sample preparation for the measurement of Ru/Os binding to DNA and proteins using ICP-MS: Binding of Ru/Os to cellular DNA and proteins was analyzed in A549 cells, using ICP-MS. A549 cells were prepared and collected using the same procedure as described above. Total DNA and protein were isolated using TRI Reagent® (Sigma–Aldrich) according to the manufacturer’s procedure and concentrations were determined spectrophotometrically by measuring absorbances (Eppendorf BioPhotometer 6131).