Epitope load identifies kidney transplant recipients at risk of allosensitization following minimization of immunosuppression

1. Download : Download high-res image (301KB)
2. Download : Download full-size image

     

Automated LC-MS method for the fast stereoselective determination of methadone in plasma.

Methadone (MTD) is a chiral drug widely used for the treatment of opioid dependence for which a rapid analytical method for the determination of each enantiomer would be advantageous. In order to improve method sensitivity and to automate the entire analytical process, a column-switching configuration has been developed. An online extraction system coupled to a cellulose-based chiral stationary phase (CSP), namely Chiralcel OJ-R, was used and detection was performed by mass spectrometry. Fifty microl of plasma were injected into the liquid chromatography-mass spectrometry (LC-MS) system after addition of acetonitrile (ACN) containing methadone deuterated D9 (MTD-D9) (internal standard) and centrifugation. For the rapid extraction step, a large particle size support was selected. A baseline separation of MTD enantiomers was obtained in less than 12 min. Trueness and precision were evaluated with control samples at 500 ng/ml of (R,S)-methadone. Trueness values were 106.6% and 103.0% for (R)-MTD and (S)-MTD, respectively, with a coefficient of variation inferior to 2.5% for both compounds. Finally, a good concordance was found using this method for analysis of plasma samples from patients in maintenance treatment as compared to a previously described HPLC-UV method after liquid-liquid extraction.     

Predictive molecular and genetic toxicology

Conclusion The search for a link between cellular and molecular events involved in delayed-type CHS reactions and the early molecular activation of xenobiotics is a new field of research. It should largely contribute to the debate on the best way forward for predictive toxicology in general.     

The Intubation Difficulty Scale (IDS): Proposal and Evaluation of a New Score Characterizing the Complexity of Endotracheal Intubation

Background: A quantitative scale of intubation difficulty would be useful for objectively comparing the complexity of endotracheal intubations. The authors have developed a quantitative score that can be used to evaluate intubating conditions and techniques with the aim of determining the relative values predictive factors of intubation difficulty and of the techniques used to decrease such difficulties.

Methods: An Intubation Difficulty Scale (IDS) was developed, based on parameters known to be associated with difficult intubation. It was then evaluated prospectively in a group of 311 consecutive prehospital intubations and 315 intubations in an operating room. In the operating room, the IDS was compared with two other parameters: the time to completion of intubation and the visual analog scale (VAS). Time was measured by an independent observer. Operators in both groups completed a checklist regarding the conditions of intubation.

Results: There is a good correlation between the IDS scale and the VAS assessment of difficulty and time to completion of intubation. VAS and time to completion have a significant but lesser correlation to each other. Comparison of IDS with operator-assessed subjective categorical impression of difficulty by Kruskall-Wallis was statistically significant.     

Allogeneic bone marrow transplantation in chronic lymphocytic leukemia: 17 cases. Report from the EBMTG

Allogeneic bone marrow transplantation (BMT) was performed in 17 patients with chronic lymphocytic leukemia (CLL): 15 resistant and two untreated forms. There were 12 males and five females with a mean age of 40 years (32-49). The conditioning regimens and graft-versus-host disease (GVHD) prophylaxis varied. Successful engraftment was obtained in 15 evaluable cases. Lymphocytosis and clinical symptoms subsided in all but one case. All 15 evaluable patients developed acute GVHD. Among the 17 patients grafted, one early death was observed at the 15th day post-BMT, and one refractory patient died 2 months after BMT. Of the remaining 15 patients in complete remission (CR), four died from GVHD, hemorrhage and graft failure, and two relapsed at 7 and 54 months after BMT and died. Nine patients are alive in CR with a mean follow-up of 25.6 months (4-48). Chimerism was complete in eight patients and partial in the two T cell-depleted cases. In one case, an immunoglobulin gene rearrangement study showed no residual disease. These results suggest that allogenic BMT might be an alternative and possible curative therapy for refractory CLL in young patients when performed relatively early in the disease.     

Rapid combined genotyping of factor V, prothrombin and methylenetetrahydrofolate reductase single nucleotide polymorphisms using minor groove binding DNA oligonucleotides (MGB probes) and real-time polymerase chain reaction.

Risk factors for cardiovascular diseases and venous thromboembolism involve both acquired and hereditary conditions. Among the latter, mutations in genes coding for coagulation factors (factor V Leiden [Arg506Gly], G20210A in the 3'-untranslated region of factor II ) and variant C677T of the methylenetetrahydrofolate reductase (MTHFR ) are often involved and co-inherited. These three factors were genotyped simultaneously in the same 96-well plate, using a real-time polymerase chain reaction (PCR) Taqman assay and minor groove binding DNA oligonucleotides (MGB probes). While primers and MGB probes matched their corresponding single nucleotide polymorphism (SNP), the real-time MGB program was identical for each target gene. Homozygous wild-type (WT; -/-), heterozygous (+/-) or homozygous (+/+) variants (n = 362) were selected for factor V (n = 115, with -/-, 40; +/-, 40; +/+, 35), factor II (n = 122, with -/-, 60; +/-, 60; +/+, 2), and MTHFR (n = 120, with -/-, 40; +/-, 40; +/+, 40), according to the results of conventional PCR-restriction fragment length polymorphism (PCR-RFLP), but the allelic discrimination was performed blind. Results of the real-time MGB and PCR-RFLP assays were identical. This new assay was easy and fast with high throughput, without risk of molecular carryover, and cost-effective for laboratories utilizing the Taqman or related fluorescence reading methods. These advantages make it particularly suitable for large-scale combined genotyping of several polymorphisms in the routine setting.     

Synthesis and characterisation of novel nanospheres made from amphiphilic perfluoroalkylthio-beta-cyclodextrins.

This work describes the synthesis of new amphiphilic perfluorohexyl- and perfluorooctyl-propanethio-beta-cyclodextrins and the comparison of the ability of these molecules and alkyl analogue, nonanethio-beta-cyclodextrin to form nanospheres. Nanospheres were prepared using nanoprecipitation method (perfluoroalkylthio-beta-cyclodextrin in THF [0.11 x 10(-3)M], stirring rate 700rpm, addition of aqueous phase at 64 degrees C into organic phase at 50 degrees C). They were characterised by Photon Correlation Spectroscopy (PCS) and by electron microscopy (SEM and cryo-TEM). The nanospheres prepared from these new beta-cyclodextrin derivatives have an average size of 260nm, and appear to be spherical in cryo-TEM images. Whereas alkyl analogue forms polydisperse aggregates with sizes in the range 60-350nm.