The validation testing for both high range assays included

The validation testing for both high range assays included precision and accuracy testing with 6 replicates of quality control samples prepared at 4 concentrations (nicotine <7.4% C.V., <7.0% bias; cotinine <11.3% C.V., <10.0% bias), recovery (nicotine 72–78%, cotinine 81–86%), multiple-lot quantitation, hemolyzed sample integrity, turbid sample integrity, aqueous stock/sub-stock solution GSK256066 at −20°C in polypropylene, long-term clinical sample stability (220 days), freeze/thaw stability (6 cycles under white light), short-term stability (49h under white light at ambient temperature), and post-preparative stability (142h). All data from the method validation was reviewed by an independent quality assurance unit as per Celerion standard operating procedures.
3-HPMA, HBMA, and CEMA in urine A bioanalytical assay developed to quantitate 3-Hydroxypropyl Mercapturic Acid (3-HPMA), Hydroxybutyl Mercapturic Acid (HBMA) and Cyanoethylmercapturic Acid (CEMA) in human urine. A 10 point standard calibrator line was established in artificial urine (CST Technologies) across the 3-HPMA and HBMA (20.0–5000ng/mL) and CEMA (0.275–207ng/mL) analytical ranges. Calibration standards, quality control samples and clinical samples were supplemented with stable label internal standards (15N13C3-3-HPMA, 15N13C3-HBMA, and 15N13C3-CEMA). The target analytes were retained through a 96-well solid-phase extraction process. The eluents were dried under a stream of nitrogen gas. The xtracts were reconstituted in deionized water and were injected onto an LC-MS/MS system for detection. Negative ions were monitored in multiple reaction monitoring (MRM) mode. The validation testing included precision and accuracy testing with 6 replicates of quality control samples prepared at 6 concentrations (3-HPMA <10.2% C.V., <6.3% bias; HBMA <7.6C.V., 13.5% bias, CEMA <10.0% C.V., <4.4% bias), recovery (3-HPMA 54–64%, HBMA 76–83%, CEMA 82–86%), multiple-lot quantitation, stock/sub-stock solution stability at −20°C in polypropylene, long-term clinical sample stability (273 days at −20°C), freeze/thaw stability (7 cycles in brown tubes), short-term stability (43h in brown tubes at ambient temperature), and post-preparative stability (152h in a polypropylene 96 well plate at 5°C). All data from the method validation was reviewed by an independent quality assurance unit as per Celerion standard operating procedures.
Aromatic amines in urine A bioanalytical assay developed to quantitate o-toluidine (o-tol), 1-aminonaphthalene (1-NA), 2-aminonaphthalene (2-NA) and 4-aminobiphenyl (4-ABP) in human urine. An eight point standard calibrator line was established in artificial urine (CST Technologies) across the o-toluidine (50.0–1000pg/mL), 1-aminonaphthalene and 2-aminonaphthalene (2.50–500pg/mL), and 4-aminobiphenyl (1.50–75.0pg/mL) analytical ranges. Calibration standards, quality control samples and clinical samples were supplemented with stable label internal standards ([13C6]o-toluidine, [13C4]2-aminonaphthalene, [13C6]4-aminobiphenyl). The target analytes were retained through a liquid/liquid extraction process. The extraction solvent was dried under a stream of nitrogen gas. The extracts were reconstituted in a polar organic solvent and injected onto an LC-MS/MS system for detection. Positive ions were monitored in multiple reaction monitoring (MRM) mode. The validation testing included precision and accuracy testing with 6 replicates of quality control samples prepared at 5 concentrations (o-toluidine <9.9% C.V., <3.7% bias; 1-NA <11.7% C.V., <3.3% bias; 2-NA <12.0C.V., <5.6% bias, 4-ABP <5.2% C.V., <8.1% bias), recovery (o-toluidine 80.0%, 1-NA 53.0–55.0%, 2-NA 81.0–84.0%, 4-ABP 82.0–84.0%), multiple-lot quantitation, stock/sub-stock solution stability at −20°C in polypropylene, long-term clinical sample stability (185 days at −20°C), freeze/thaw stability (6 cycles at ambient temperature in brown polypropylene tubes), short-term stability (54h at ambient temperature), and post-preparative stability (337h). All data from the method validation was reviewed by an independent quality assurance unit as per Celerion standard operating procedures.