[Recommendations for the control of documents and the establishment of a documentary system]. / Recommandations pour la maîtrise des documents et la constitution d'un système documentaire.

The quality management system that must be implemented in a MBL to meet the requirements of the standard NF EN ISO 15189 is based, among other things, on the creation and use by staff of a documentary system approved and updated. This documentary system is constituted by external documents (standards, suppliers' documents...) and internal documents (quality manual, procedures, instructions, technical and quality recordings...). A procedure of the documentary system control must be formalized. The documentary system should be modeled in order to identify the various procedures to be drafted and the incurred risks in the case a document would be missing in this system. Each document must be indexed in a unique way and document management must be carried out rigorously. The use of document management software is a great help to manage the life cycle of documents.

[Hygiene and security management in medical biology laboratory]. / Hygiène et sécurité au laboratoire de biologie médicale.

Risk management in Medical Biology Laboratory (MBL) which includes hygiene and waste management, is an integrated process to the whole MBL organisation. It is composed of three stages: risks factors identification, grading and prioritization, and their evaluation in the system. From the legislation and NF EN ISO 15189 standard's requirements viewpoint, prevention and protection actions to implement are described, at premises level, but also at work station environment's one (human resources and equipments) towards biological, chemical, linked to gas, to ionizing or non ionizing radiations and fire riks, in order not to compromise patients safety, employees safety, and quality results. Then, although NF EN 15189 standard only enacts requirements in terms of prevention, curative actions after established blood or chemical exposure accident are defined.

[Recommendations for waste management]. / Recommandations pour la gestion des déchets.

Laboratory waste management must ensure the safety of patients and staff, limiting the environmental impacts and control waste disposal budget. Sorting of waste must be carried out at the source. The packaging must be adapted, allowing easy identification of specific disposal routes. With regard to wastes for human or animal health care and/or related research (DASRI), packages must comply with the regulations, standards and ADR if necessary. Storage provisions differ according to the amount of DASRI produced. Waste collection is carried out directly on the place of activity by a certified service provider. Non pre-treated DASRI is incinerated in specific approved plants for a T ° > 1,200 °C. Special provisions also exist for chemical waste and radioactive waste, the latter being regulated by ANDRA.

Hair analysis by LC-MS as evidence of nalbuphine abuse by a nurse.

Individuals in any profession can succumb to chemical abuse. Among the healthcare profession, nurses represent a specific group because of their ease of access to drugs, particularly narcotics. Opioids, potentially highly addictive agents, are usually their drug of choice. Nalbuphine, a synthetic opioid analgesic, is prescribed for moderate-to-severe acute pain, for chronic pain syndromes, and in obstetrics to decrease the adverse respiratory effect of opioid epidural administration. The case of a nurse who was suspected of drug misuse after the disappearance of two nalbuphine ampules in an obstetrics service is described. Because of discrepancies in the results of her blood and urine samples, a sample of head hair was subsequently collected from the nurse. A hair analysis of nalbuphine by liquid chromatography-mass spectrometry has not been previously described. Following decontamination and grinding, hair was mixed with a Söerensen buffer, then subjected to ultrasonic treatment (1 h), and extracted with ethyl acetate. A quantitative analysis was performed with two channels (30 and 45 V), and it is based on a m/z 358 for nalbuphine and a m/z 330 for methylclonazepam as an internal standard. The method was linear from 0.020 to 12 ng/mg of hair (R(2) = 0.972), and the limit of detection and limit of quantitation are 0.020 ng/mg. Accuracy (CV), assessed at 0.4 and 1.6 ng/mg of hair, was 6.18% and 5.77%, respectively, for intraday assays and 4.5% and 10.9% for interday assays. Recovery efficiency at 1.6 ng/mg and 8 ng/mg of hair was 100% and 97.4%, respectively. The hair specimen from the nurse (6 cm) was cut into three equal lengths. Nalbuphine, venlafaxine, and nordiazepam were detected. The concentration of nalbuphine was similar in the three hair locks: 5.07, 7.06, and 5.70 ng/mg of hair. A hair analysis revealed the repeated intake of nalbuphine by the nurse. This person was treated for depression for several months with Effexor (venlafaxine) and Nordaz (nordiazepam) prior to the investigation. Hair appears to be a unique matrix to provide evidence for chronic drug exposure by establishing a historic record that is not possible by blood or urine analysis.

[Serial determination of crimidine by HPLC/SE/SM in a patient ingesting a "mouse trap"]. / Détermination sérique de la crimidine par CLHP/ES/SM chez un patient ayant ingere un "souricide foudroyant".

Crimidine (2 chloro, 4 methyl, 6 dimethyl amidopyrine) is a synthetic rodenticide which causes acute poisonings after oral ingestion in human. Major toxic effects are consciousness disorders, hypertonic coma and convulsions. Toxic level in human is about 5 mg/Kg. An intoxication case is reported. Five serums collected at different times were analyzed with HPLC/ES/MS. Crimidine was extracted with ethylacetate with recovery over 80%. Linearity was up to 800 micrograms/L. LOQ and LOD were 0.5 and 0.3 microgram/L respectively. The coefficients of variation were less than 10% for repeatability and reproductibility. Serum levels varied from 368 micrograms/L for H0 to 64 micrograms/L for H10 and elimination of crimidine was linear in time.

Determination serique de la crimidine par clhp/es/sm chez un patient ayant ingere un « souricide foudroyant ¼.

Crimidine (2 chloro, 4 methyl, 6 dimethyl amidopyrine) is a synthetic rodenticide which causes acute poisonings after oral ingestion in human. Major toxic effects are consciousness disorders, hypertonic coma and convulsions. Toxic level in human is about 5 mg/Kg. An intoxication case is reported. Five serums collected at different times were analyzed with HPLC/ ES/MS. Crimidine was extracted with ethylacetate with recovery over 80 %. Linearity was up to 800 µg/L. LOQ and LOD were 0.5 and 0.3 µg/L respectively. The coefficients of variation were less than 10 % for repeatability and reproductibility. Serum levels varied from 368 µg/L for H0 to 64 µg/L for H10 and elimination of crimidine was linear in time.

Separation and quantification of paraquat and diquat in serum and urine by capillary electrophoresis.

The use of capillary electrophoresis (CE) for simultaneous qualitative and quantitative detection of paraquat (PQ) and diquat (DQ) in both serum and urine was investigated. The two herbicides were extracted from biological fluids with liquefied phenol. Serum required a deproteinization with chloroform and ammonium sulfate as pretreatment. The extracts were hydrodynamically injected and the complete separation was carried out in 10 min, using a capillary tube (75 microm i.d., 500 mm) of fused silica containing 50 mM phosphate buffer (pH 2.50) as the carrier. UV absorbance detection at 200 nm was performed by an on-column detector. The analytes were characterized by their respective migration times. Analytical recoveries were 52.6% for PQ and 62.6% for DQ in serum, and 71.4% and 59.3%, respectively, in urine. The linearity was studied up to 4 mg/L and the limits of detection (LODs) were better than 5 pg/mL in serum or urine. The CE method described was applied to the characterization of two lethal poisonings and results were related.

Colchicine poisoning: report of a fatal case with body fluid and post-mortem tissue analysis by high-performance liquid chromatography.

A case involving a suicide by the ingestion of colchicine tablets is presented. Liquid chromatography has been used to measure the drug level in blood and in post-mortem tissues of the patient (a 42-year-old man). Plasma concentration 24 h after ingestion was 4.5 ng/mL. On autopsy, the kidney showed the highest concentration (396 ng/g). High concentrations were also found in the liver (347 ng/g) and heart (334 ng/g). Low concentrations were detected in the lung (58 ng/g), muscle (10 ng/g) and brain (5 ng/g).

Genotypic and phenotypic analysis of the polymorphic thiopurine S-methyltransferase gene (TPMT) in a European population.

1. Characterization of allelic variants of the TPMT gene (TPMT) responsible for changes in TPMT activity, and elucidation of the mechanism by which these alleles act, are required because of the clinical importance of this polymorphism for patients receiving thiopurine drugs. 2. We defined the mutational and allelic spectrum of TPMT in a group of 191 Europeans. Using PCR-SSCP, we screened for mutation the entire coding sequence, the exon-intron boundaries, the promoter region and the 3'-flanking region of the gene. Six mutations were detected throughout the ten exons and seven TPMT alleles were characterized. Four of them, TPMT*2, *3A, *3C and *7, harbouring the known mutations, G238C, G460A, A719G or T681G, were nonfunctional and accounted for 0.5, 5.7, 0.8 and 0.3% of the allele totality, respectively. 3. Within the promoter region, six alleles corresponding to a variable number of tandem repeats (VNTR), were identified. VNTR*V4 and *V5a which harbour four or five repeats of a 17-18 bp unit, were the most frequent (55% and 34%, respectively). The other VNTR alleles, having from five to eight repeats, were rarer. 4. The TPMT phenotype was correctly predicted by genotyping for 87% of individuals. A clear negative correlation between the total number of repeats from both alleles and the TPMT activity level was observed, indicating that VNTRs contribute to interindividual variations of TPMT activity. Therefore, additional analysis of the promoter region of TPMT can improve the phenotype prediction rate by genotyping.

Detection of known and new mutations in the thiopurine S-methyltransferase gene by single-strand conformation polymorphism analysis.

To detect mutations in the thiopurine S-methyltransferase gene (TPMT), we have developed a strategy based on single-strand conformation polymorphism (SSCP) analysis of the gene amplified by polymerase chain reaction (PCR). The sensitivity of the method was first evaluated by analyzing DNA samples from five individuals, including two high methylators (HMs), two intermediate methylators (IMs), and one deficient methylator (DM). TPMT alleles and mutations in each of these individuals had previously been characterized by conventional PCR-based assays and direct sequencing analysis. All mutations were associated with particular shifts in the electrophoretic mobility of DNA fragments, allowing their identification. We further tested the efficiency of the strategy to detect new TPMT mutations. For this purpose, additional DNAs from 15 IMs and 15 HMs were submitted to PCR-SSCP analysis. A total of 7 alleles were characterized, including two new alleles. The first one, termed TPMT*1A, harbors a single mutation C-->T at nucleotide -178 in exon 1 and was detected in a HM subject. The second one, termed TPMT*7, was characterized by a T-->G transversion at nucleotide 681 in exon 10. This allele should be a nonfunctional allele of the TPMT gene since it was observed in combination with a wild-type allele in an intermediate methylator. We conclude that the PCR-SSCP strategy we developed could be advantageously used to fully characterize the extent of allelic variation at the TPMT gene locus in populations and thus to improve our understanding of the genetic polymorphism of TPMT activity, which has considerable consequences for the toxicity and efficacy of therapeutically important and widely used drugs.

Congenital bilateral absence of the vas deferens (CBAVD) and cystic fibrosis transmembrane regulator (CFTR): correlation between genotype and phenotype.

To assess better the link between congenital bilateral absence of the vas deferens (CBAVD) and cystic fibrosis (CF), we compared sweat chloride values, analysis of the CFTR intron 8 poly(T) tract length and analysis of 10 exons in a population of 38 patients with CBAVD. The data indicate that this population can be divided into three groups of patients. In the first group of 15 patients with abnormal sweat chloride (> 60 mmol/l), the frequency of CF mutations is high. In the second group of 18 patients with equivocal sweat chloride (between 40 and 60 mmol/l), the frequency of the 5T variant is high; 6 patients have a delta F508 mutation and a 5T variant and 1 patient is homozygous for the 5T variant; a 5T variant has been detected in 3 other patients, and a delta F508 mutation in another patient. A third group of 5 patients is probably not related to CF: these patients have other congenital abnormalities of the urogenital tract, low chloride values (< 40 mmol/l) and apparently no abnormality of the CF gene.