Icpms 2030

The ICP-MS (Shimadzu ICP-MS 2030, Shimadzu, Kyoto, Japan) technique, equipped with a collision cell with a helium flow of 6 mL/min, was used to quantify the silver concentration (silver ion and AgNPs) in the liver tissue and blood samples. All collected biological materials were pre-digested for 4 h at 80 °C using a Thermomixer (Thermomixer comfort, Eppendorf SE, Hamburg, Germany) in 150 µL of HNO3 (1%) (Merck Suprapure, Merck, Darmstadt, Germany). Prior to analysis, the cooled samples were diluted 100 times in 1% HNO₃. Measurements were performed on 107Ag and 103Rh as internal standards, using the method of external calibration in three replications. The results were presented in the form of a concentration unit. The concentration was reported based on the calibration curve carry-out for silver standards (Silver and Zinc Standards for ICP, Sigma-Aldrich, Merck, Germany) in a concentration range of 0—25 µg/L.

LA-ICP-MS was performed as described previously.^{14 (link)} Aneurysms were cut into 10 µm cryosections at −20°C and mounted on SuperFrost Plus adhesion slides (Thermo Scientific). The LA-ICP-MS analysis was performed using an LSX 213 G2+ laser system (CETAC Technologies, Omaha, USA) which was equipped with a 2-volume HelEx II cell connected via Tygon tubing to an ICPMS-2030 (Shimadzu, Kyoto, Japan). With a spot size of 10 µm samples were ablated via line-by-line scan using a scan speed of 30 µm/s and 800 mL/min He as transport gas. The analysis was performed in collision gas mode with He as collision gas, with 50 ms integration time for the analyzed isotopes ^{31 (link)}P, ⁵⁷Fe, and ⁶⁴Zn and 75 ms integration time for the 2 Gd isotopes ¹⁵⁸Gd and ¹⁶⁰Gd. Matrix-matched standards based on gelatin were used for the quantification of Gd. Nine gelatin standards (10% w/w) including a blank, were spiked with different Gd concentrations (1 to 500 µg/g). Averaged intensities of the scanned lines of the standards were in good linear correlation with a regression coefficient R² = 0.998 within this concentration range. Limit of detection (LOD) and limit of quantification (LOQ), calculated with the 3σ- and 10σ-criteria, were 16 ng/g and 54 ng/g Gd. The quantification and visualization were performed using in-house developed software (WWU Münster, Münster, Germany).

All Ag⁺ solutions were prepared in deionized water with no additional modifications. The exact Ag⁺ concentration in the solutions was quantified by ICP-MS analysis. Here and in all other analyses, metal quantification was performed on Shimadzu ICP-MS 2030 (Kyoto, Japan) with scandium (⁴⁵Sc) as an internal standard. The quantification was performed taking into account the ¹⁰⁷Ag isotope. The 1% HNO₃ was utilized for the dilution. Instead, the bLTF suspension was prepared in deionized water with concentration of 5 mg/mL (by weight) and adjusted to pH 5.0. The pH adjustment was performed with 1% HNO₃ or 0.1 NaOH. For the study, bLTF standard purchased from Sigma-Aldrich (Steinheim, Germany) was utilized. Before the study, the physicochemical characterization of the bLTF was performed; among others, the iron content was determined. According to obtained data, the utilized bLTF was in holo-form. The results are presented in the article by O. Pryshchepa et al. (2022) [22 (link)]. The bLTF from the same batch was utilized in both studies.

Tomato leaves and roots were washed and collected in liquid nitrogen after 10 d Cd treatment. Samples were put in the oven for 30 min at 115 °C then transferred to 60 °C for total dryness. The dried sample was ground and mixed with an acid solution that contained HClO₄ and HNO₃ (1:3, v:v). The samples were digested at 180 °C and the solution was then evaporated to 1-2 mL for dilution. The remained solution was diluted with deionized water to a final volume of 50 mL and inductively coupled plasma mass spectrometry (ICPMS-2030, Shimadzu Company, Kyoto, Japan) was used for determining Cd content [25 (link)].
For Cd localization, the Leadmium^TM Green AM probe (Invitrogen, Carlsbad, CA, USA) was used for Cd staining according to the manufacturer’s instructions. The root tips were stained by immersing in dye solution for 3 h in dark at room temperature; the sample was then washed three times with buffer (0.85% NaCl). A Nikon A1 confocal microscope (Nikon, Tokyo, Japan) was used to detect the Cd localization, and the excitation/emission wavelengths of GFP were 488 nm/510–530 nm [41 (link)]. The mean fluorescence intensity values of Cd stained root tips were detected by ImageJ 1.53 analysis software (National Institute of Health, Bellevue, WA, USA) and relative fluorescence intensity normalized to the intensity of the wild-type group under Cd stress.

The minerals’ composition was determined according to Serrano-Díaz, Sánchez, Martínez-Tomé, Winterhalter and Alonso [4 (link)], with slight modifications. Freeze-dried bread samples weighing 0.5 g were digested with 10 mL of 65% HNO₃ (v/v) using a microwave reactor digestor (CEM Mars one, Matthews, NC, USA) for 30 min with a temperature ramp whose final temperature was 200 °C. All samples were filtered (Whatman qualitative filter paper 90 mm), diluted with ultrapure deionized water 1:50 (v/v), and stored at 4 °C. Total concentrations of macronutrients (Ca, Mg, Na, and K) and micronutrients (Zn, Cu, Mn, and Fe) in the previously mineralized samples were quantified with an Inductively Coupled Plasma Mass Spectrometer (ICPMS-2030, Shimadzu, Kyoto, Japan). Internal standards included calcium (44Ca), magnesium (26Mg), sodium (23Na), potassium (39K), zinc (66Zn), copper (65Cu), manganese (55Mn), and iron (56Fe), and the calibration curves used for quantification showed good linearity (R² ≥ 0.998). The results were expressed as mg/100 g of dw (dry weight).

For elemental bioimaging, LA-ICP-MS was performed using an LSX-213-G2+ laser system (CETAC Technologies, USA), equipped with a two-volume HelExII cell connected via Tygon tubing to an ICP-MS spectrometer (ICPMS-2030, Shimadzu, Japan). Quantification and visualization were performed with an in-house developed software (Robin Schmid, WWU Münster, Germany). Histological samples were ablated via line-by-line scan with a spot size of 15 µm, a scan speed of 45 µm/s, and 800 mL/min helium as carrier gas. Subsequent analysis was conducted in collision gas mode using He as collision gas and 100 ms integration time for the analyzed isotopes ³¹P, ⁶⁴Zn, ¹⁵⁸Gd, and ¹⁶⁰Gd. For quantification of Gd, matrix-matched gelatin-based standards were used. Nine gelatin standards (10% w/w), including a blank, were spiked with different Gd concentrations ranging from 1 to 5.000 µg/g. There was good linear correlation for averaged intensities of scanned lines with a regression coefficient R² = 0.9999 over this concentration range. Limit of detection (LOD) and limit of quantification (LOQ), calculated with the 3σ- and 10σ-criteria, were 8 ng/g and 28 ng/g Gd, respectively.