Detection of anti-SARS-CoV-2 antibodies in dried blood spots utilizing manual or automated spot extraction and electrochemiluminescence immunoassay (ECLIA).

Serological test methods to detect anti-SARS-CoV-2 antibodies represent a major measure to manage the pandemic caused by the coronavirus disease 2019 (COVID-19). In this communication, test results obtained from minimal-invasively collected dried blood spot (DBS) specimens, which can be sampled 'at home' without the need of medically trained personnel, are compared to conventionally collected venous blood samples. DBS samples were prepared for analysis either manually or by a card extraction robot, and electrochemiluminescence assay (ECLIA) characteristics, assay readout values as well as stability data covering a period of more than 200 days are provided. Constant anti-SARS-CoV-2 antibody readouts of quality control DBS were obtained over the entire test period using DBS specimens stored under dry and dark conditions. In addition, test results obtained from individuals tested twice within 10 months post-infection indicated a consistent presence of antibodies.

Simultaneous quantification of THC-COOH, OH-THC, and further cannabinoids in human hair by gas chromatography-tandem mass spectrometry with electron ionization applying automated sample preparation.

The detection of Δ9 -tetrahydrocannabinol (THC), cannabidiol (CBD) and cannabinol (CBN) in hair, for the purpose of identifying cannabis consumption, is conducted in many forensic laboratories. Since external contamination of hair with these cannabis components cannot be excluded, even after hair decontamination, only the detection of THC metabolites such as 11-nor-9-carboxy-Δ9 -tetrahydrocannabinol (THC-COOH) or 11-hydroxy-Δ9 -tetrahydrocannabinol (OH-THC), is considered to prove cannabis consumption. At present, testing for THC metabolites is not standard practice due to its analytical complexity. For these reasons, we developed a novel method for the detection of THC-COOH and OH-THC as well as THC, CBD, and CBN in one single analytical run using gas chromatography-tandem mass spectrometry (GC-MS/MS) with electron ionization. After manual hair washing and grinding, sample preparation was fully automated, by means of a robotic autosampler. The hair extraction took place by digestion with sodium hydroxide. A solid-phase extraction (SPE) was chosen for sample clean-up, using a mixed-mode anion exchange sorbent. Derivatization of all analytes was by silylation. The method has been fully validated according to guidelines of the Society of Toxicological and Forensic Chemistry (GTFCh), with a limit of detection (LOD) of 0.2 pg/mg for THC-COOH and OH-THC and 2 pg/mg for THC, CBD and CBN, respectively, thus fulfilling the Society of Hair Testing (SoHT) recommendations. The validated method has been successfully applied to our routine forensic case work and a summary of data from authentic hair samples is given, as well as data from proficiency tests.

Automated GC-MS Determination of Δ9-Tetrahydrocannabinol, Cannabinol and Cannabidiol in Hair.

The determination of Δ(9)-tetrahydrocannabinol (THC), cannabinol (CBN) and cannabidiol (CBD) in hair is a major routine task in forensic laboratories worldwide. A comprehensively automated liquid-liquid extraction (LLE) method has been developed. The automation was carried out by an x-y-z sample robot equipped with modules capable of shaking, centrifugation and solvent evaporation. It comprises digestion of hair in sodium hydroxide solution, LLE, extract evaporation, reconstitution in silylation reagent, inlet derivatization and GC-MS analysis. Method validation guidelines of the Society for Toxicological and Forensic Chemistry were fulfilled. The limit of quantification (LOQ) was 0.01 ng/mg for THC, 0.06 ng/mg for CBN and 0.03 ng/mg for CBD. This is below the required LOQ for THC (0.02 ng/mg) in medical psychological assessments in Germany. Also it is far below the required LOQ of the Society of Hair Testing of 0.1 ng/mg for THC. Four-round robin tests were passed successfully and several post- and ante-mortem samples were analyzed. To date the method is routinely employed at the Institute of Legal Medicine in Giessen, Germany. To the best of our knowledge, this is the first publication on a comprehensively automated classical LLE workflow in the field of hair analysis.

Development and validation of an automated liquid-liquid extraction GC/MS method for the determination of THC, 11-OH-THC, and free THC-carboxylic acid (THC-COOH) from blood serum.

The analysis of Δ(9)-tetrahydrocannabinol (THC) and its metabolites 11-hydroxy-Δ(9)-tetrahydrocannabinol (11-OH-THC), and 11-nor-9-carboxy-Δ(9)-tetrahydrocannabinol (THC-COOH) from blood serum is a routine task in forensic toxicology laboratories. For examination of consumption habits, the concentration of the phase I metabolite THC-COOH is used. Recommendations for interpretation of analysis values in medical-psychological assessments (regranting of driver's licenses, Germany) include threshold values for the free, unconjugated THC-COOH. Using a fully automated two-step liquid-liquid extraction, THC, 11-OH-THC, and free, unconjugated THC-COOH were extracted from blood serum, silylated with N-methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA), and analyzed by GC/MS. The automation was carried out by an x-y-z sample robot equipped with modules for shaking, centrifugation, and solvent evaporation. This method was based on a previously developed manual sample preparation method. Validation guidelines of the Society of Toxicological and Forensic Chemistry (GTFCh) were fulfilled for both methods, at which the focus of this article is the automated one. Limits of detection and quantification for THC were 0.3 and 0.6 µg/L, for 11-OH-THC were 0.1 and 0.8 µg/L, and for THC-COOH were 0.3 and 1.1 µg/L, when extracting only 0.5 mL of blood serum. Therefore, the required limit of quantification for THC of 1 µg/L in driving under the influence of cannabis cases in Germany (and other countries) can be reached and the method can be employed in that context. Real and external control samples were analyzed, and a round robin test was passed successfully. To date, the method is employed in the Institute of Legal Medicine in Giessen, Germany, in daily routine. Automation helps in avoiding errors during sample preparation and reduces the workload of the laboratory personnel. Due to its flexibility, the analysis system can be employed for other liquid-liquid extractions as well. To the best of our knowledge, this is the first publication on a comprehensively automated classical liquid-liquid extraction workflow in the field of forensic toxicological analysis. Graphical abstract GC/MS with MPS Dual Head at the Institute of Legal Medicine, Giessen, Germany. Modules from left to right: (quick) Mix (for LLE), wash station, tray 1 (vials for extracts), solvent reservoir, (m) VAP (for extract evaporation), Solvent Filling Station (solvent supply), cooled tray 2 (vials for serum samples), and centrifuge (for phase separation).

Comprehensive automation of the solid phase extraction gas chromatographic mass spectrometric analysis (SPE-GC/MS) of opioids, cocaine, and metabolites from serum and other matrices.

The analysis of opioids, cocaine, and metabolites from blood serum is a routine task in forensic laboratories. Commonly, the employed methods include many manual or partly automated steps like protein precipitation, dilution, solid phase extraction, evaporation, and derivatization preceding a gas chromatography (GC)/mass spectrometry (MS) or liquid chromatography (LC)/MS analysis. In this study, a comprehensively automated method was developed from a validated, partly automated routine method. This was possible by replicating method parameters on the automated system. Only marginal optimization of parameters was necessary. The automation relying on an x-y-z robot after manual protein precipitation includes the solid phase extraction, evaporation of the eluate, derivatization (silylation with N-methyl-N-trimethylsilyltrifluoroacetamide, MSTFA), and injection into a GC/MS. A quantitative analysis of almost 170 authentic serum samples and more than 50 authentic samples of other matrices like urine, different tissues, and heart blood on cocaine, benzoylecgonine, methadone, morphine, codeine, 6-monoacetylmorphine, dihydrocodeine, and 7-aminoflunitrazepam was conducted with both methods proving that the analytical results are equivalent even near the limits of quantification (low ng/ml range). To our best knowledge, this application is the first one reported in the literature employing this sample preparation system.

Increasing productivity for the analysis of trace contaminants in food by gas chromatography-mass spectrometry using automated liner exchange, backflushing and heart-cutting.

Laboratories focusing on residue analysis in food are continuously seeking to increase sample throughput by minimizing sample preparation. Generic sample extraction methods such as QuEChERS lack selectivity and consequently extracts are not free from non-volatile material that contaminates the analytical system. Co-extracted matrix constituents interfere with target analytes, even if highly sensitive and selective GC-MS/MS is used. A number of GC approaches are described that can be used to increase laboratory productivity. These techniques include automated inlet liner exchange and column backflushing for preservation of the performance of the analytical system and heart-cutting two-dimensional GC for increasing sensitivity and selectivity. The application of these tools is illustrated by the analysis of pesticides in vegetables and fruits, PCBs in milk powder and coplanar PCBs in fish. It is demonstrated that considerable increase in productivity can be achieved by decreasing instrument down-time, while analytical performance is equal or better compared to conventional trace contaminant analysis.

Derivatisation and gas chromatography-chemical ionisation mass spectrometry of selected synthetic and natural endocrine disruptive chemicals.

Methods for ultra trace detection of endocrine disruptive chemicals (EDCs) are needed because of their low levels of impact. Twenty-one EDCs were selected, including 17beta-estradiol, 17alpha-ethinylestradiol, 17beta-testosterone and bisphenol A. Derivatisation with eight different fluorine containing compounds was examined. All EDCs could be derivatised automatedly (autosampler) with heptafluorobutyric acid (HFB) anhydride and trifluoroacetic acid (TFA) anhydride, respectively. The detection of these HFB and TFA derivatives in different chemical ionisation modes was studied. Fourteen different reagent gases, including methane, ammonia, acetone and water, were tested with the HFB and TFA derivatives in the negative chemical ionisation mode. Furthermore both types of derivatives were measured in positive chemical ionisation mode. Methane or water provide a good detection of all 21 TFA derivatives and create mass spectra with few fragmentation and characteristic mass peaks. This could serve as a basis for tandem or multiple mass spectrometric measurements.