Creatinine parameter assay kit

F₂-Isoprostanes are prostaglandin-like compounds produced as a result of free radical lipid peroxidation of arachidonic acid [22 (link)]. Levels of urinary F₂-isoprostanes were determined by the liquid chromatography-mass spectrometry (LC/MS/MS) assay method developed by Davies et al [23 (link)]. Urinary creatinine concentrations were determined using the Creatinine Parameter Assay kit (R&D Systems Inc., Minneapolis, MN, USA) and following the manufacture’s guidelines. We expressed F₂-isoprostane levels relative to urinary creatinine levels to normalize for excretion rate.

Clean-catch midstream urine was collected from each patient in a 50 mL sterile container in the morning. For biopsy-concurrent aLN patients, urine samples were procured within 5 days before kidney biopsy. Urine samples were aliquoted to avoid repeated freeze-thaw cycles and stored at − 80°C. Urinary L-selectin (uL-selectin) was assayed using a commercially available human L-selectin enzyme-linked immunosorbent assay (ELISA) kit (DY728, R&D System; ELH-LSelectin, Raybiotech; 1:20) according to the manufacturer’s instruction. uL-selectin was normalized by urine creatinine using Creatinine Parameter Assay Kit (KGE005, R&D Systems).

Urine was collected from individual mice every 2 weeks from 8 weeks of age for over 16 h in metabolic cages (Shinano Manufacturing Co., Tokyo, Japan). The concentration of urine albumin and creatinine were measured using DC Protein Assay Reagent (Bio-Rad Laboratories, Hercules, CA, USA) and Creatinine Parameter Assay Kit (R&D Systems, Minneapolis, MN, USA), respectively, and the urine albumin-creatinine ratio was quantified.
Mouse serum anti-dsDNA Ab was measured using an enzyme-linked immunosorbent assay (ELISA) (Shibayagi, Shibukawa, Japan). Human serum anti-TRIM21 Ab was measured using an ELISA kit (Dr. Fooke Laboratorien, Neuss, Germany), according to the manufacturer's protocols.

Resveratrol, L-(+)-tartaric acid, quercetin, (+)-catechin hydrate sulfatase (Helix pomatia, Type H-1, sulfatase ≥ 10,000 units/g), and β-glucuronidase (Helix pomatia, Type HP-2, ≥100,000 units/mL) were purchased from Sigma-Aldrich (St. Louis, MO, USA). 8-iso prostaglandin F2α and 8-iso prostaglandin F2α-d4 were purchased from Cayman Chemical Company (Ann Arbor, MI, USA). (±)-Tartaric-2,3-d2 acid was purchased from C/D/N Isotopes (QC, Pointe-Claire, QC, Canada). Fisetin and formic acid (LC-MS grade) were obtained from TCI chemicals (Portland, OR, USA). LC-MS grade water and acetonitrile were purchased from Sigma-Aldrich (St. Louis, MO, USA). A creatinine parameter assay kit was purchased from R&D systems (Minneapolis, MN, USA), and Millex-GV Filter (0.22 µm) was obtained from EMD Millipore (Burlington, MA, USA).

The samples of plasma, sputum, and urine were collected at enrollment and stored at −70°C. Levels of EDN in plasma, supernatant of sputum, and urine were measured using K‐EDN kit (SKIMS‐BIO Co., Seoul, Korea) as previously described.²¹ Serum and urine levels of leukotriene E4 (LTE₄) were analyzed by liquid chromatography–tandem mass spectrometry. LTE₄‐d5 (Cayman Chemical Company, Ann Arbor, MI, USA) was used as a deuterated internal standard. Chromatographic separations were performed using the Waters Acquity UPLC system (Waters) with a Hypersil GOLD column (2.1 × 100 mm, 1.9 μm: ThermoFisher Scientific, San Jose, CA, USA) on a concentration gradient. Data acquisition was performed using an API5500 triple quadrupole mass spectrometer (AB Sciex, Framingham, MA, USA) equipped with an Electrospray ionization source. For the quantitative determination of creatinine in urine samples, 10 μl of urine sample was applied to the Creatinine Parameter Assay Kit (R&D Systems, Minneapolis, MN, USA).