Diagnostic performance of line-immunoassay based algorithms for incident HIV-1 infection

Background: Serologic testing algorithms for recent HIV seroconversion (STARHS) provide important information for HIV surveillance. We have previously demonstrated that a patient's antibody reaction pattern in a confirmatory line immunoassay (INNO-LIA™ HIV I/II Score) provides information on the duration of infection, which is unaffected by clinical, immunological and viral variables. In this report we have set out to determine the diagnostic performance of Inno-Lia algorithms for identifying incident infections in patients with known duration of infection and evaluated the algorithms in annual cohorts of HIV notifications. Methods: Diagnostic sensitivity was determined in 527 treatment-naive patients infected for up to 12 months. Specificity was determined in 740 patients infected for longer than 12 months. Plasma was tested by Inno-Lia and classified as either incident (< = 12 m) or older infection by 26 different algorithms. Incident infection rates (IIR) were calculated based on diagnostic sensitivity and specificity of each algorithm and the rule that the total of incident results is the sum of true-incident and false-incident results, which can be calculated by means of the pre-determined sensitivity and specificity. Results: The 10 best algorithms had a mean raw sensitivity of 59.4% and a mean specificity of 95.1%. Adjustment for overrepresentation of patients in the first quarter year of infection further reduced the sensitivity. In the preferred model, the mean adjusted sensitivity was 37.4%. Application of the 10 best algorithms to four annual cohorts of HIV-1 notifications totalling 2'595 patients yielded a mean IIR of 0.35 in 2005/6 (baseline) and of 0.45, 0.42 and 0.35 in 2008, 2009 and 2010, respectively. The increase between baseline and 2008 and the ensuing decreases were highly significant. Other adjustment models yielded different absolute IIR, although the relative changes between the cohorts were identical for all models Conclusions: The method can be used for comparing IIR in annual cohorts of HIV notifications. The use of several different algorithms in combination, each with its own sensitivity and specificity to detect incident infection, is advisable as this reduces the impact of individual imperfections stemming primarily from relatively low sensitivities and sampling bias

Capsaicin-Induced Changes in LTP in the Lateral Amygdala Are Mediated by TRPV1

The transient receptor potential vanilloid type 1 (TRPV1) channel is a well recognized polymodal signal detector that is activated by painful stimuli such as capsaicin. Here, we show that TRPV1 is expressed in the lateral nucleus of the amygdala (LA). Despite the fact that the central amygdala displays the highest neuronal density, the highest density of TRPV1 labeled neurons was found within the nuclei of the basolateral complex of the amygdala. Capsaicin specifically changed the magnitude of long-term potentiation (LTP) in the LA in brain slices of mice depending on the anesthetic (ether, isoflurane) used before euthanasia. After ether anesthesia, capsaicin had a suppressive effect on LA-LTP both in patch clamp and in extracellular recordings. The capsaicin-induced reduction of LTP was completely blocked by the nitric oxide synthase (NOS) inhibitor L-NAME and was absent in neuronal NOS as well as in TRPV1 deficient mice. The specific antagonist of cannabinoid receptor type 1 (CB1), AM 251, was also able to reduce the inhibitory effect of capsaicin on LA-LTP, suggesting that stimulation of TRPV1 provokes the generation of anandamide in the brain which seems to inhibit NO synthesis. After isoflurane anesthesia before euthanasia capsaicin caused a TRPV1-mediated increase in the magnitude of LA-LTP. Therefore, our results also indicate that the appropriate choice of the anesthetics used is an important consideration when brain plasticity and the action of endovanilloids will be evaluated. In summary, our results demonstrate that TRPV1 may be involved in the amygdala control of learning mechanisms

Real-Time Investigation of the H/D Exchange Kinetics of Porphyrins and Oligopeptides by Means of Neutral Cluster-Induced Desorption/Ionization Mass Spectrometry

The kinetics of the H/D exchange reaction in angiotensin II, hexaglycine (Gly₆), Co­(II)­tetra­(3-carboxyphenyl)­porphyrin, and tetra­(4-carboxyphenyl)­porphyrin were followed in real time by mass spectrometry employing desorption/ionization induced by neutral SO₂ clusters. The change of the isotope patterns with increasing degree of deuteration was recorded as a function of D₂O exposure and the underlying H/D exchange kinetics, i.e., the dependence of the different degrees of deuteration on time, were deduced. The results were modeled by means of Monte Carlo simulations taking into account different reaction constants for the H/D exchange reaction at different functional groups. In the case of the investigated porphyrins, the rate constants were directly assigned to the functional groups involved; in the case of the peptides, reaction at the explicit functional groups and the backbone chain of the molecules could be discriminated

Targeted Small-Molecule Identification Using Heartcutting Liquid Chromatography-Infrared Ion Spectroscopy

Infrared ion spectroscopy (IRIS) can be used to identify molecular structures detected in mass spectrometry (MS) experiments and has potential applications in a wide range of analytical fields. However, MS-based approaches are often combined with orthogonal separation techniques, in many cases liquid chromatography (LC). The direct coupling of LC and IRIS is challenging due to the mismatching timescales of the two technologies: an IRIS experiment typically takes several minutes, whereas an LC fraction typically elutes in several seconds. To resolve this discrepancy, we present a heartcutting LC-IRIS approach using a setup consisting of two switching valves and two sample loops as an alternative to direct online LC-IRIS coupling. We show that this automated setup enables us to record multiple IR spectra for two LC-features from a single injection without degrading the LC-separation performance. We demonstrate the setup for application in drug metabolism research by recording six m/z-selective IR spectra for two drug metabolites from a single 2 μL sample of cell incubation extract. Additionally, we measure the IR spectra of two closely eluting diastereomeric biomarkers for the inborn error of metabolism pyridoxine-dependent epilepsy (PDE-ALDH7A1), which shows that the heartcutting LC-IRIS setup has good sensitivity (requiring ∼μL injections of ∼μM samples) and that the separation between closely eluting isomers is maintained. We envision applications in a range of research fields, where the identification of molecular structures detected by LC–MS is required