Between-run Relative Standard Deviations Research Articles

Determination of flurbiprofen in rat plasma using ultra-high performance liquid chromatography-tandem mass spectrometry and its application in a pharmacokinetic study.

A rapid and sensitive ultra-high performance liquid chromatography-tandem mass spectrometry method was developed to determine flurbiprofen in rat plasma. A triple quadrupole tandem mass spectrometer equipped with an electrospray ionization (ESI) source was used in negative ion mode. Acetonitrile precipitation was selected to prepare samples. Flurbiprofen and internal standard flurbiprofen-d5 were analyzed on an Acquity UPLC BEH C18 column with the mobile phase consisting of acetonitrile and water, and a gradient procedure was used for separation. The retention time of flurbiprofen was 0.67 min, and the whole running time was only 1.2 min. The detection was performed on a triple quadrupole tandem mass spectrometer using multiple reaction monitoring mode via an ESI source with optimized mass spectrometry parameters. The calibration curve was linear in the range of 25.0-1.00 × 104 ng/mL (r ≥ 0.99). The within-run and between-run relative standard deviations were not more than 13.9%. The within-run and between-run relative errors were from -9.0% to 3.4%. There was no significant matrix effect, and recovery was high. This method was fully validated, including whole blood stability in rat plasma, and successfully applied to the pharmacokinetic study in which 100% incurred sample reanalysis met the criteria.

Determination of six arsenic species in urine by high-performance liquid chromatography-inductively coupled plasma-mass spectrometr

Objective: Establishing an analysis method for the detection of six arsenic compounds trivalent arsenic, pentavalent arsenic, methyl arsenic, dimethyl arsenic, arsenical choline and arsenical betaine in urine by high-performance liquid chromatography-inductively coupled plasma-mass spectrometry (HPLC-ICP-MS) and providing a theoretical basis for health assessment of arsenic poisoning patients. Methods: HPLC-ICP-MS was used to detect six arsenic compounds with the hamilton PRP-X100 column and aqueous ammonium phosphate in a gradient elution as mobile phase. Results: There was a good linear relationship in the selected range of six arsenic compounds evidenced by a linear correlation coefficient greater than 0.999. The detection limit of 6 arsenic compounds was 0.46~0.57 μg/L, and the minimum quantitative concentration were 1.52~1.91 μg/L. The recovery rates of six arsenic compounds were 96.5%~106.4%, The within-run and between-run relative standard deviation was 2.9%~6.2% and 3.8%~8.7%. Conclusion: The established method with high accuracy and precision is suitable for detecting arsenic levels in urine of occupational arsenic exposure population.

Graphite furnace atomic absorption spectrometry for the determination of trace gallium in whole blood

Objective: To establish a graphite furnace atomic absorption spectrometry method for the determination of trace gallium in whole blood. Methods: From January to May 2021, the five factors of ashing temperature, ashing time, atomization temperature, atomization time and matrix modifier concentration in the determination of gallium in whole blood by graphite furnace atomic absorption spectrometry were optimized by using L(16) (4(5)) orthogonal test design. At the same time, within-run, between-run, spiking recovery test and other methodological indicators were tested. Results: Under the optimized detection conditions, the linear range of determination of gallium in whole blood by graphite furnace atomic absorption spectrometry was 0.29-100.00 μg/L (r=0.9991) . The within-run and between-run relative standard deviations (RSD) of repetitive measurement at 10.0, 50.0, 80.0 μg/L concentration levels were 2.3%-4.4% and 1.5%-3.6%, the recovery rate of spiking was 98.1%-103.8%, and the detection limit of the method was 0.13 μg/L. Conclusion: Graphite furnace atomic absorption spectrometry for the determination of trace gallium in whole blood is easy to operate, has a wide linear range, low detection limit, accurate and reliable results, which is suitable for occupational health examinations and the determination of acute gallium poisoning.

Research and development of an air sampling tube for chlorobenzene compounds and its supporting determination method

Objective: To establish a GDH-3 air sample tube for simultaneous determination of twelve kinds of chlorobenzene compounds (CBs) in workplace air by gas chromatography. And to established a matching determination method. Methods: In October 2020, the vapor and aerosol CBs in workplace air were collected by GDH-3 air sampling tube, and desorption and elution with 3.00 ml toluene for 15 min, then the solution separated by DB-23 capillary column, and finally detected with microcell electron capture detector. Results: The quantitative determination ranges of twelve isomers of CBs were 0.71×10(-3)-2000.00 mg/L, with the correlative coefficients were 0.99967-0.99998. The minimum detectable concentrations were 0.04-112.63 μg/m(3), and the minimum quantification concentrations were 0.14-375.42 μg/m(3) (15.00 L sample, 3.00 ml sample solution) . The average elution efficiencies were 96.00%-104.00%. The within-run relative standard deviations (RSDs) were 2.54%-6.12%, and the between-run RSDs were 3.85%-7.87%. Sealed samples could be stable at room temperature for at least 15 days. Conclusion: GDH-3 air sample tube can be used for simultaneous determination of twelve kinds of CBs in workplace air by gas chromatography. The established supporting measurement method meets the measurement requirements of the occupational health standard detection method, and the it's suitable for the simultaneous determination of 12 kinds of CBS in the air.

Determination of arsenic species compound in blood by high performance liquid chromatographyatomic fluorescence spectrometry

Objective: To establish a method for determination of arsenic species in blood with high performance liquid chromatography-atomic fluorescence spectrometry (HPLC-AFS) . Methods: The effect of mobile phase about chromatography separation and sample pretreatment conditions and atomic fluorescence spectrometry detection parameters has been optimized to reliably measure the following four kinds of species arsenic compound including arsenic[As (III) ]、dimethylarsinic acid (DMA) 、monomethylarsonic acid (MMA) and arsenate[As (V) ] in acute intoxication human blood. The method of technical standard about within-run, between-run and recoveries of standard were optimized. Results: The method showed As (III) linear relationship was 2.63-100.00 μg/L, The detection limit was 2.63 μg/L. The relative coefficient (r) was 0.9999; DMA linear relationship was 3.21-100.00 μg/L, The detection limit was 3.21 μg/L. The r was 0.9992; MMA linear relationship was 3.41-100.00 μg/L, The detection limit was 3.41 μg/L. The r was 0.9998; As (V) linear relationship was 3.90-100.00 μg/L, The detection limit was 3.90 μg/L. The r was 0.9996. The average recovery of four species arsenic in tested samples ranged from 91.3%-99.8% with the relative standard deviation (RSD) from 2.39% to 4.05%. The within-run and between-run relative standard deviations (RSD) of repetitive measurement at 10.00, 40.00, 80.00 μg/L concentration levels were 1.99%-4.59% and 2.72%-4.53%. Conclusion: This method is low detection limit, good accurate and high sensitivity, proposed method had been applied to the analysis of arsenic species in blood samples those who acute intoxication or poisoning diagnosis.

Determination of trace bismuth by atomic fluorescence spectrometry in blood

Objective: To establish a method for determination trace bismuth in blood by atomic fluorescence spectrometry (AFS) . Methods: 4.0 ml nitric acid and 1.0 ml perchloric acid was added into 1.0 ml blood sample then through automation graphite digestion instrument digested, after that 1.0 ml thiocarbamide-vitamin (10%) was injected, 8% HCl constant volume to 10.0 ml, the bismuth was detected by atomic fluorescence spectrometry with 5.0 ml digestive sample. Results: The method showed a linear relationship within the range of 0.4-50.0 μg/L (r=0.999 7) . The within-run and between-run relative standard deviations (RSD) of repetitive measurement at 10.0, 20.0, 40.0 μg/L concentration levels were 2.2%-4.9% and 3.0%-4.0%. The detection limit was 0.032 μg/L. The recoveries of bismuth were 93.0%-103.9%. Conclusion: This method is low detection limit, good accurate and high sensitivity. It has been applied for determination of trace bismuth in blood samples those who need occupation health examination or poisoning diagnosis.

Determination of trace gallium by graphite furnace atomic absorption spectrometry in urine

To establish a method for determination trace gallium in urine by graphite furnace atomic absorption spectrometry (GFAAS). The ammonium dihydrogen phosphate was matrix modifier. The temperature effect about pyrolysis (Tpyr) and atomization temperature were optimized for determination of trace gallium. The method of technical standard about within-run, between-run and recoveries of standard were optimized. The method showed a linear relationship within the range of 0.20~80.00 μg/L (r=0.998). The within-run and between-run relative standard deviations (RSD) of repetitive measurement at 5.0, 10.0, 20.0 μg/L concentration levels were 2.1%~5.5% and 2.3%~3.0%. The detection limit was 0.06 μg/L. The recoveries of gallium were 98.2%~101.1%. This method is simple, low detection limit, accurate, reliable and reproducible. It has been applied for determination of trace gallium in urine samples those who need occupation health examination or poisoning diagnosis.

Dispersive liquid-liquid microextraction based on solidification of floating organic drop and high-performance liquid chromatography to the analysis of cocaine's major adulterants in human urine.

A simple method has been proposed for the determination of cocaine's major adulterants (caffeine, levamisole, lidocaine, phenacetin, diltiazem, and hydroxyzine) in human urine by dispersive liquid-liquid microextraction based on solidification of floating organic drop (DLLME-SFO) in combination with high-performance liquid chromatography - photodiode array detector (HPLC-PDA). The reversed-phase chromatographic separation was obtained with a column C18 extended (250×4.6mm; 5μm; 80Å) in gradient elution mode using acetonitrile-trifluoroacetic acid 0.026% (v,v) (pH=2.5) at 1mLmin-1 as mobile phase, at 25°C, and detection at 235nm. The analysis time was 25min. This condition had the best resolution factors (>1.15), retention factors (>0.68), number of plates (>2094.9), and separation factors (>1.05) for all targets, indicating a good separation. The kind of extraction and dispersive solvent were investigated for unifactorial design. The buffer pH, the volume of extraction and disperser solvent, and the amount of salt were optimized for full factorial design. Under optimum conditions, human urine samples were alkalized with 0.5M sodium phosphate buffer (pH 10) and added to sodium chloride (20%m/v). Acetonitrile (150μL) and 1-dodecanol (30μL) were used as dispersive and extraction solvent, respectively. The method presented linear range of 312.5-3125ngmL-1 to caffeine and levamisole and 187.5-1875ngmL-1 to lidocaine, phenacetin, diltiazem, and hydroxyzine. The limit of quantification was 187.5ngmL-1 to lidocaine, phenacetin, diltiazem, and hydroxyzine and 312.5ngmL-1 for caffeine and levamisole. The recovery mean values were between 6.0 and 42.6%. The method showed good precision and accuracy, with within- and between-run relative standard deviation and relative error less than 15%. The samples were stable after freeze-thaw cycle and short-term room temperature stability tests. Besides, this method was satisfactorily applied in urine of cocaine users. It is expected that this method, which was the first to combine the use of DLLME-SFO and HPLC-PDA for the determination of cocaine's major adulterants in human urine, will contribute to the accuracy in the diagnosis of acute intoxication, the proper planning of therapeutic measures, as well as to the favorable prognostic of cocaine intoxicated patients.

Simultaneous quantitation of polymyxin B1, polymyxin B2 and polymyxin B1-1 in human plasma and treated human urine using solid phase extraction and liquid chromatography-tandem mass spectrometry.

Two liquid chromatographic-tandem mass spectrometric (LC-MS/MS) methods have been developed and validated for the quantitative determination of polymyxin B1, polymyxin B2 and polymyxin B1-1 concentrations in human plasma and treated urine. During method development, technical challenges such as the separation of structural isomers polymyxin B1and polymyxin B1-1 and nonspecific binding in urine samples were encountered and overcome. Two automated solid phase extraction methods were used to extract plasma samples (100μL) and urine samples (200μL) and the resulting extracts were analyzed using reversed phase LC-MS/MS with an electrospray (ESI) interface and selected reaction monitoring (SRM) in the positive ionization mode. Both methods were validated over a calibration curve range of 5.00-2000ng/mL with a linear regression and 1/x(2) weighting. The between-run relative standard deviation (%RSD) ranged from 4.5 to 9.5% for the plasma assay and from 1.1 to 7.1% for the urine assay. For the plasma assay, the between-run accuracy ranged from 100.5 to 115.2% of nominal at all QC concentrations including the LLOQ. For the urine assay, the between-run accuracy ranged from 92.0 to 106% of nominal at all QC concentrations including the LLOQ. The extraction recoveries for all polymyxins in both assays were between 54.0 and 64.2%. Long term matrix storage stability for all polymyxins was established at both -20°C and -70°C for up to 85 days in human plasma and for up to 55 days in treated human urine. Both assays were used for the measurement of polymyxin B1, polymyxin B2 and polymyxin B1-1 concentrations in human plasma and treated urine for the determination of bioequivalence and toxicokinetic parameters in clinical studies.

Development and Validation of an HPLC-UV Assay to Quantify Plasma Levels of Sulfametrol: A Preferential Antibiotic in Children.

Sulfonamides in combination with trimethoprim are frequently used antibiotics. They work synergistic. In infections with Pneumocystis jiroveci or Stenotrophomonas maltophilia, higher dosages are indicated than in other infections. Therapeutic drug monitoring (TDM) is warranted to assure the efficacy while limiting toxicity. Although trimethoprim in combination with sulfamethoxazole is the most common combination with established TDM target concentrations, the intravenous formulation is not suited for children because of its additives ethanol and propylene glycol to increase solubility. An alternative can be sulfametrol in combination with trimethoprim. When sulfametrol was introduced in the hospital, there was a need for a TDM method for sulfametrol. A High Pressure Liquid Chromatography-Ultraviolet (HPLC-UV) detection method for sulfametrol determination in plasma was developed and validated according to the International Conference on Harmonization guidelines. Linearity, limit of detection, lower limit of quantification, recovery, process efficiency, selectivity, within-run precision, between-run precision, and sample stability were tested. All tested parameters met the required criteria. For linearity, r was 0.9948, lower limit of quantification was 10 mg/L, and limit of detection was 6 mg/L. Recovery was 100.4% and process efficiency 94.4%. Selectivity was met with no interfering peaks at the retention time of 4.2 minutes. Between-run precision and within-run precision were evaluated by replicating quality control levels, resulting in a within-run relative average standard deviation of 0.8% and a between-run relative standard deviation of 2.3%. Recovery of the samples after storing 8 days was 101.9% and recovery of already tested vials was 98.8% after 48 hours. In conclusion, an HPLC-UV method for sulfametrol determination in human plasma was developed and validated. The method is fast, accurate, reproducible, and has a short analysis time. It is now being used in routine TDM in our clinic.

Dual dispersive liquid–liquid microextraction for determination of phenylpropenes in oils by gas chromatography–mass spectrometry

A novel, simple and quick sample preparation method was developed and used for pre-concentration and extraction of six phenylpropenes, including anethole, estragole, eugenol, methyl eugenol, safrole and myristicin, from oil samples by dual dispersive liquid–liquid microextraction. Gas chromatography-mass spectrometry was used for determination and separation of compounds. Several experimental parameters affecting extraction efficiency were evaluated and optimized, including forward-extractant type and volume, surfactant type and concentration, water volume, and back-extractant type and volume. For all analytes (10–1000ng/mL), the limits of detection (S/N≧3) ranged from 1.0 to 3.0ng/mL; the limits of quantification (S/N≧10) ranged from 2.5 to 10.0ng/mL; and enrichment factors ranged from 3.2 to 37.1 times. Within-run and between-run relative standard deviations (n=6) were less than 2.61% and less than 4.33%, respectively. Linearity was excellent with determination coefficients (r2) above 0.9977. The experiments showed that the proposed method is a simple, effective, and environmentally friendly method of analyzing phenylpropenes in oil samples.

Development and validation of LC/ESI-MS method for the detection and quantification of exogenous ceramide NP in stratum corneum and other layers of the skin

Ceramides (CERs) are integral parts of the intercellular lipid lamellae of the stratum corneum (SC), which provide the barrier function of the skin. Administration of CERs deep into the SC may help to restore the barrier function in affected or aged skin. However, quantification of the amount of CER penetrated into the target site needs a selective and sensitive analytical method. Therefore, an LC/ESI-MS method was developed and validated for the detection and quantification of exogenous CER [NP] in the SC as well as other skin layers. The strategy involved synthesis of ceramide [NP]-D3-18 (deuterated CER [NP]) to distinguish it from the endogenous CER [NP] in MS on weight basis. The method was linear over 10–800 ng/ml and sensitive with a limit of detection (LOD) and limit of quantification (LOQ) of 3 and 10 ng/ml, respectively. It was also accurate with within-run and between-run percentage recoveries of 97.1–103.2 and 99.0–104.9, respectively. The within-run and between-run relative standard deviations (RSDs) were 0.9–5.4% and 2.1–7.4%, respectively, suggesting the method is precise. The method was highly selective and the matrix effect was too minimal with matrix factor (MF) mean and RSD values of 1.002 and 4.57%, respectively.

Serum/plasma methylmercury determination by isotope dilution gas chromatography-inductively coupled plasma mass spectrometry

A method for the determination of methylmercury in plasma and serum samples was developed. The method uses isotope dilution with 198Hg-labeled methylmercury, extraction into dichloromethane, back-extraction into water, aqueous-phase ethylation, purge and trap collection, thermal desorption, separation by gas chromatography, and mercury isotope specific detection by inductively coupled plasma mass spectrometry. By spiking 2 mL sample with 1.2 ng tracer, measurements in a concentration interval of (0.007–2.9) μg L −1 could be performed with uncertainty amplification factors <2. A limit of quantification of 0.03 μg L −1 was estimated at 10 times the standard deviation of concentrations measured in preparation blanks. Within- and between-run relative standard deviations were <10% at added concentration levels of 0.14 μg L −1, 0.35 μg L −1 and 2.8 μg L −1, with recoveries in the range 82–110%. Application of the method to 50 plasma/serum samples yielded a median (mean; range) concentration of methylmercury of 0.081 (0.091; <0.03–0.19) μg L −1. This is the first time methylmercury has been directly measured in this kind of specimen, and is therefore the first estimate of a reference range.

Rapid and direct measurement of free concentrations of highly protein‐bound fluoxetine and its metabolite norfluoxetine in plasma

Fluoxetine (F) and its active N-demethylated metabolite, norfluoxetine (NF), are selective serotonin re-uptake inhibitors that bind extensively to plasma proteins. Development and validation of a novel method for measuring free concentrations of F and NF in plasma are reported here. The plasma filtrate was prepared by a high-speed short-duration ultrafiltration (UF) and then submitted directly to a short-column liquid chromatography/tandem mass spectrometric (LC/MS/MS) assay. There was no significant matrix effect on the analysis, and non-specific binding of the analytes to the UF devices was negligible. For validation of the method, the recovery of the free analytes was compared to that from an optimized equilibrium dialysis method, and analyte stability was examined under conditions mimicking the sample storage, handling, and analysis procedures. The linearity range was 0.37-12 ng/mL for F and NF; the within-run and between-run relative standard deviations were less than 11.9%, and accuracies across the assay range were 100 +/- 10.3%. This new method was then further validated in a pharmacokinetic (PK) study in beagle dogs receiving a single oral dose of fluoxetine hydrochloride. The integrity of the resulting PK data of free F and NF was absolute. The PK data indicate that the novel method is accurate and reliable. To our knowledge this is the first report describing a rapid and reliable method for direct measurement of free concentrations of F and NF in plasma, which will be useful for clinical pharmacokinetic/pharmacodynamic studies of F. Furthermore, the strategies described herein may be applied to the development and validation of methods for measuring the free concentrations of other drugs in plasma.

Effective Use of Liquid Chromatography-Mass Spectrometry (LC/MS) in the Routine Clinical Laboratory for Monitoring Sirolimus, Tacrolimus, and Cyclosporine

The authors describe an isocratic liquid chromatographic/electrospray single quadrupole mass spectrometric assay suitable for routine therapeutic monitoring of the antirejection drugs tacrolimus, sirolimus, and cyclosporine in blood. The drugs and added internal standards, ascomycin, desmethoxysirolimus, and cyclosporine G, are extracted from a protein-free supernatant of patient blood on a Strata SDB-L styrene-divinylbenzene polymeric extraction cartridge (Phenomenex, Torrance, CA). A Gilson Aspec XL-4 (Gilson Instruments, Middleton, WI) is programmed to perform the extraction and transfer the extract to autosampler vials. The extract is automatically injected onto a C18 column held at 75 degrees C. Mobile phase, acetonitrile/water 90/10 (vol/vol), is pumped at 0.5 mL/min through a silica saturating column positioned in the column oven before the injector valve. Detection is by selected ion monitoring of the sodium adduct of each analyte, [M + 23]. A linear response is achieved for tacrolimus and sirolimus from limit of quantitation (LOQ) 1.0 microg/L to at least 80.0 microg/L. Cyclosporine was linear from LOQ 25 microg/L to at least 2000 microg/L. Interferences and ionization suppression are minimal. Analytic recovery for cyclosporine is 99.9 +/- 6.2% over a range of 104-1162 microg/L; sirolimus 107.7 +/- 9.3% over a range of 3.2-29.2 microg/L; and tacrolimus 101.9 +/- 2.5% over a range of 2.9-38.1 microg/L. Between-run relative standard deviations are 3.0% to 5.1% over a range of 9.8 microg/L to 30.7 microg/L for tacrolimus; 5.5% to 7.6% over a range of 13.5 microg/L to 35.2 microg/L for sirolimus; and 2.6% to 4.3% over a range of 186.0 microg/L to 1428.7 microg/L for cyclosporine. In the authors' laboratory, the method is robust and shows improved selectivity, precision, detection limits, and lower cost per test over available immunoassays. The authors find the method suitable for routine monitoring and therapy management of these drugs with over 50,000 samples analyzed over the course of 1 year.

Rapid determination of tafenoquine in small volume human plasma samples by high-performance liquid chromatography–tandem mass spectrometry

A method was developed for the determination of tafenoquine ( I) in human plasma using high-performance liquid chromatography–tandem mass spectrometry. Prior to analysis, the protein in plasma samples was precipitated with methanol containing [ 2H 3 15N]tafenoquine ( II) to act as an internal standard. The supernatant was injected onto a Genesis-C 18 column without any further clean-up. The mass spectrometer was operated in the positive ion mode, employing a heat assisted nebulisation, electrospray interface. Ions were detected in multiple reaction monitoring mode. The assay required 50 μl of plasma and was precise and accurate within the range 2 to 500 ng/ml. The average within-run and between-run relative standard deviations were <7% at 2 ng/ml and greater concentrations. The average accuracy of validation standards was generally within ±4% of the nominal concentration. There was no evidence of instability of I in human plasma following three complete freeze–thaw cycles and samples can safely be stored for at least 8 months at approximately −70°C. The method was very robust and has been successfully applied to the analysis of clinical samples from patients and healthy volunteers dosed with I.

Evaluation of accuracy and uncertainty of ELISA assays for the determination of interleukin-4, interleukin-5, interferon-gamma and tumor necrosis factor-alpha.

The comparison of analytical results calls for validation of the assays used, i.e. for the documentation of accuracy (trueness and precision), linearity and specificity. In addition, there is a growing demand for evaluation and documentation of traceability and uncertainty of analytical results. However, models for establishing the traceability and uncertainty of immunoassay results are lacking. Sandwich enzyme-linked immunosorbent assays (ELISAs) were developed for determination of the human cytokines interleukin-4 (IL-4), interleukin-5 (IL-5), interferon-y (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha). The accuracy of each of the assays was evaluated in the ranges of 1-15 microg/l (IL-4), 0.001-1 microg/l (IL-5), 0.5-2.5 microg/l (IFN-T) and 0.14-2.2 microg/l (TNF-alpha). Other evaluated performance characteristics were the limit of detection (LOD), immunological specificity, and robustness. Traceability was ensured by the use of World Health Organization International Standards (WHO IS). An uncertainty budget, which combined the contribution from all known uncertainty components, was established for each cytokine ELISA. The between-run relative analytical standard deviation (RSDA) of the assessed ELISAs was found to be in the range of 11-18%, except for IL-5 where RSDA increased at decreasing concentrations. The LOD was 0.12 microg/l, 0.0077 microg/l, 0.0069 microg/l and 0.0063 microg/l for IL-4, IL-5, IFN-gamma and TNF-alpha, respectively. Traceability to the WHO IS was established for each of the cytokines. The combined relative standard uncertainty (U(result)/C(result)) was 28%, 22-62%, 28% and 24% for the IL-4, IL-5, IFN-gamma and TNF-alpha results, respectively. The major contributions to uncertainty came from the relative analytical standard deviation and from the uncertainty of the mass concentration of the WHO IS. The uncertainty of the WHO IS was not stated in the accompanying certificate and was evaluated by other means. The largest sources of uncertainty were located outside our laboratory. This means that the possibilities to improve the reliability of the results produced by the ELISAs are very limited. The task of evaluating measurement uncertainty would be much easier if producers of international reference standards reported the uncertainty of the value of standards. The model for evaluating uncertainty presented in this paper is applicable to other types of assays and to most analytical methods.

A novel combination of immunoreaction and ICP-MS as a hyphenated technique for the determination of thyroid-stimulating hormone (TSH) in human serum

The novel coupling method based on the hyphenation of immunoreaction with ICP-MS was developed and applied to the determination of thyroid-stimulating hormone (TSH) in human serum. In this system, TSH was firstly captured by anti-TSH monoclonal antibodies immobilized on a solid support. Biotinylated anti-TSH monoclonal antibodies and Eu3+-labelled streptavidin were then added to form the complex, captured antibody–TSH–biotinylated anti-TSH antibody–Eu3+-labelled streptavidin. After unbound reactants were washed away, Eu3+ bound to the complex was extracted by 1% HNO3 and measured by ICP-MS. The Eu signal was directly proportional to the serum TSH concentration. The limit of detection was 0.5 mIU L−1 and the assay was linear up to 170 mIU L−1. The within-run (n = 6) and between-run (n = 6) relative standard deviations (RSD) for low, medium and high concentration standards did not exceed 10%. The TSH results obtained by this procedure correlated well with those obtained by radioimmunoassay (r2 = 0.9639).

Determination of plasma heparin by micellar electrokinetic capillary chromatography

The micellar electrokinetic capillary chromatography (MECC) method is reported for the separation of heparin, and for the possibility of direct determination of free heparin in plasma. The conditions for MECC were: pH 8.5, 25 mM sodium dodecyl sulfate (SDS), 25 mM borate buffer, with a 30 cm×50 μm ID fused-silica capillary. The sample was detected with a UV-detector at 270 nm with heparin as external standard. The recovery rate was 95.6–98.7%. This method was linear in the range 80–7000 U l −1. The within-run and between-run relative standard deviations were lower than 3.1 and 4.5%, respectively. It is suggested that this MECC method may be used to determine blood samples containing high levels of heparin.

Automated Extraction of Lysergic Acid Diethylamide (LSD) and N-demethyl-LSD from Blood, Serum, Plasma, and Urine Samples Using the Zymark RapidTrace™ with LC/MS/MS Confirmation

A forensic procedure for the quantitative confirmation of lysergic acid diethylamide (LSD) and the qualitative confirmation of its metabolite, N-demethyl-LSD, in blood, serum, plasma, and urine samples is presented. The Zymark RapidTrace was used to perform fully automated solid-phase extractions of all specimen types. After extract evaporation, confirmations were performed using liquid chromatography (LC) followed by positive electrospray ionization (ESI+) mass spectrometry/mass spectrometry (MS/MS) without derivatization. Quantitation of LSD was accomplished using LSD-d3 as an internal standard. The limit of quantitation (LOQ) for LSD was 0.05 ng/mL. The limit of detection (LOD) for both LSD and N-demethyl-LSD was 0.025 ng/mL. The recovery of LSD was greater than 95% at levels of 0.1 ng/mL and 2.0 ng/mL. For LSD at 1.0 ng/mL, the within-run and between-run (different day) relative standard deviation (RSD) was 2.2% and 4.4%, respectively.