Diagnostic accuracy of a new fluoroenzyme immunoassay for the detection of TSH receptor autoantibodies in Graves' disease

Purpose: Thyrotropin receptor (TSHR) autoantibodies (TRAbs) are a hallmark of Graves’ disease (GD). The aim of this study was to evaluate the diagnostic accuracy of a new third generation automatic fluorescence enzyme immunoassay for TRAb measurement in GD, in comparison with two current IMAs. Methods: Sera of 439 subjects (57 patients with untreated GD, 34 with treated GD, 15 with GD and Graves’ orbitopathy, 52 with multinodular non-toxic goiter, 86 with Hashimoto’s thyroiditis, 20 with toxic adenoma or toxic multinodular goiter, 55 with non-thyroid autoimmune diseases and 120 normal controls) were tested for TRAbs with the ELiA™ anti-TSH-R assay (ThermoFischer Scientific, Uppsala, Sweden), the TRAK™ RIA, Brahms (Thermo Scientific, Hennigsdorf, Germany) and the Immulite™ TSI assay (Siemens Healthcare, Llanberis, UK). Results: Sensitivity and specificity of the ELiA™ anti-TSH-R assay, TRAK™ RIA and Immulite™ TSI assay were 94.7% and 99.6, 100 and 98.2%, 100 and 98.2%, respectively. Spearman’s coefficient and Passing-Bablok regression showed a satisfactory correlation between EliA™ and TRAK™ [rho: 0.925; 95% CI: 0.883-0-953. Intercept: − 0.875 (95% CI: − 2.411 to 0.194); slope: 1.086 (95% CI: 0.941 to 1.248)], and between ELiA™ and TSI™ [rho: 0.947; 95% CI: 0.912 0.969. intercept: 1.085 (95% CI: 0.665 to 2.116); slope 1.315 (95% CI:1.116 to 1.700)]. Conclusions: The diagnostic performance of ELiA™-TSH-R assay is comparable to that of some current TRAb assays. It may be adopted into clinical practice for the differential diagnosis of hyperthyroidism, to screen for transient hyperthyroidism, and to monitor disease activity and treatment effects. © 2018, The Author(s)

Laboratory Kinetic Studies of OH and CO2 Relevant to Upper Atmospheric Radiation Balance

The purpose of this project was to quantify the rates of two processes which are crucial to our understanding of radiative energy balance in the upper atmosphere. The first process is radiative emission from vibrationally hot OH radicals following the H + O3 reaction in the upper mesosphere. The importance of this process depends strongly on the OH radiative emission coefficients. Our goal was to measure the OH permanent dipole moment in excited vibrational states and to use these measurements to construct an improved OH dipole moment function and improved radiative emission coefficients. Significant progress was made on these experiments including the construction of a supersonic jet source for vibrationally excited OH radicals. Unfortunately, our efforts to transport the OH radicals into a second lower pressure vacuum chamber were not successful, and we were unable to make improved dipole moment measurements for OH. The second key kinetic process which we attempted to quantify during this project is the rate of relaxation of bend-excited CO2 by oxygen atoms. Since excitation of the bending vibrational mode of CO2 is the major cooling mechanism in the upper mesosphere/lower thermosphere, the cooling rate of this region depends crucially on the rate of energy transfer out of this state. It is believed that the most efficient transfer mechanism is via atomic oxygen but the rate for this process has not been directly measured in the laboratory at appropriate temperatures and even the room temperature rate remains controversial. We attempted to directly measure the relaxation rate Of CO2 (010) by oxygen atoms using the discharge flow technique. This experiment was set up at Aerodyne Research. Again, significant progress was achieved in this experiment. A hot CO2 source was set up, bend excited CO2 was detected and the rate of relaxation of bend excited CO2 by He atoms was measured. Unfortunately, the project ran out of time before the oxygen atom kinetic studies could be implemented

Polarization of TH2 response is decreased during pregnancy in systemic lupus erythematosus

This study evaluated some cytokines involved in the Th1-Th2 shift during pregnancy in patients with systemic lupus erythematosus (SLE) and healthy women. Twenty-seven consecutive successful pregnancies in 26 SLE patients and 28 pregnancies in 28 matched healthy subjects, as controls, were enrolled and prospectively studied. Sera obtained at first and third trimesters of pregnancy were tested for IL-1α, IL-1β, IL-2, IL-6, IL-8, IL-10, IL-12p70, INF-γ, and TNF-α with a highly sensitive, multiplexed sandwich ELISA (SearchLight Human Inflammatory Cytokine Array). Statistics were performed by SPSS package. IL-8 serum levels were higher in the first (P<0.0001) and third (P=0.003) trimesters of pregnancy in SLE patients compared with controls, INF-γ serum levels in the third trimester (P=0.009), and IL-10 serum levels in the first and third trimesters (P=0.055 and P<0.0001, respectively). IL-2 (r=0.524 P=0.010), IL-12 (r=0.549 P=0.007), IFN-γ (r=0.492 P=0.017), and IL-6 (r=0.515 P=0.020) serum levels correlated with disease activity in SLE patients in the first trimester of pregnancy. Cytokine profile was similar in patients with and without lupus nephritis both in the first and in the third trimesters of pregnancy. IL-8 serum levels were lower in patients with a previous diagnosis of antiphospholipid antibody syndrome compared with those without, both in the first and in the third trimesters of pregnancy. In SLE patients, a lower than expected decrease in Th1 cytokine serum levels was observed in the third trimester of gestation which could contribute to a lower Th2 cytokine polarization during pregnancy

Laboratory Studies of Chemical and Photochemical Processes Relevant to Stratospheric Ozone

The purpose of this project is to reduce the uncertainty in several key gas-phase kinetic processes which impact our understanding of stratospheric ozone. The main emphasis of this work is on measuring rate coefficients and product channels for reactions of HO(x) and NO(x) species in the temperature range 200 K to 240 K relevant to the lower stratosphere. The results of these studies will improve models of stratospheric ozone chemistry and predictions of perturbations due to human influences. The second year's effort has focussed the design and construction of the proposed high pressure flow reactor on three separate areas: (1) the construction of the high pressure flow reactor; (2) characterization of the turbulent flow profile; and (3) demonstration of the instrument by measuring HO2 + NO2 and HO2 + NO reaction rate coefficients

Detection of serum anti-B/B' UsnRNP antibodies in patients with connective tissue diseases by immunoblotting

Objective: To investigate the reliability of the immunoblot method in the detection of serum immunoreactivity towards the B/B' polypeptides of U small nuclear ribonucleoproteins (UsnRNP) and to assess the significance of these antibodies in connective tissue disease (CTD) patients. Methods: We tested the sera of 348 patients with CTD (101 SLE, 51 systemic sclerosis, 53 primary Sjogren's syndrome, 27 poly/dermatomyositis, 15 rheumatoid arthritis and 101 overlap CTD), of 31 matched healthy subjects and 13 patients with primary Epstein-Barr virus (EBV) infection with high titre IgG anti-EBV antibodies. IgG anti-UsnRNP antibodies were determined by immunoblotting on nuclear extract from Raji cells (an EBV-immortalised human B lymphoid cell line) and Jurkat cells (a human T lymphoid cell line). Anti-dsDNA antibodies were detected by indirect immunofluorescence on Crithidia luciliae and anti-ENA by counterimmunoelectrophoresis. Anti-dsDNA activity and avidity were measured in SLE sera by ELISA with Scatchard analysis. Results were statistically analysed by chi-square and Mann-Whitney tests. Results: A high frequency of anti-B/B' antibodies was found in the sera of CTD patients, confined to SLE (54.4%) and overlap CTD with SLE features (55,2%). Anti-B/B' immune reactivity was closely associated with other anti-UsnRNP specificities, gel precipitating anti-nRNP and anti-P antibodies. Nine out of 15 (60%) anti-B/B' positive/anti-ENA negative lupus sera on Raji blots were confirmed to be positive also on Jurkat blots. The sera from patients with EBV infection provided, on Raji blots, completely different band patterns from those obtained with auto-immune sera. Conclusions. The Sm B/B' proteins are the predominant or, at least, the most frequently targeted antigens of the UsnRNP auto-immune response in SLE and "lupus-like" overlap CTD. Moreover, anti-B/B' is diagnostically specific for CTD with SLE features. Immunoblotting on human B lymphoid cells is a reliable method, in terms of sensitivity and specificity, for the detection of anti-Sm B/B' antibodies

Producción de BeNToglucerasa para el tratamiento no invasivo de la enfermedad de Gaucher

La enfermedad de Gaucher es una enfermedad rara de almacenamiento lisosomal provocada por mutaciones en el gen que codifica para la proteína beta-glucocerebrosidasa que cataliza la hidrólisis del glicolípido glucocerebrósido a glucosa y ceramida. Las personas que padecen dicha enfermedad son incapaces de descomponer el glucocerebrósido, por lo que permanece almacenado dentro de los lisosomas.El sindrome de Gaucher se clasifica en tres tipos, dos de los cuales, el tipo II o subaguda y tipo III crónica, son de carácter neuropático y además son menos frecuentes.Actualmente las terapias están dirigidas a eliminación de sustrato o terapias de reemplazo enzimático, siendo esta última la que más éxito reporta. Se suministra la enzima exógenamente vía intravenosa y reduce algunos síntomas como la hepatoesplenomegalia. Por otro lado, presenta algunos inconvenientes, como la incapacidad de atravesar la barrera hematoencefálica, lo que la hace ineficaz para los tipos neuropáticos. Además, su administración ha de ser continua y frecuente por su corta vida media en sangre.Bionaturis pretende conseguir la administración del fármaco vía oral y atravesar la barrera hematoencefálica, a través de la acumulación de distintas versiones de glucocerebrosidasa humana mejorada utilizando la plataforma FLYLIFE y secuencias estabilizadoras y promotoras del cruce de barreras biológicas.Partiendo de la síntesis de secuencias de nucleótidos de las distintas versiones de glucocerebrosidasa, procedimos a clonarlas en vectores de expresión baculovíricos, que se utilizaron para insertar los constructos en el Master Viral DNA de Bionaturis y crear baculovirus recombinantes de trabajo o Working Viral Banks (WVB). Para ello se realizó la co-transfección de células de insectos (sf21) y se obtuvieron los baculovirus con la secuencia de interés por  recombinación homóloga. Tras la amplificación de los stocks de baculovirus, procedimos a la caracterización de los WVBs, mediante tests de identidad, pureza y potencia que incluye el análisis de la expresión del producto de interés en células de insecto. Los resultados han sido positivos y muestran la presencia de glucocerebrosidasa recombinante en sedimento celular pero no en el sobrenadante de los cultivos infectados. Nuestros siguientes pasos van  enfocados al test de expresión en larvas de lepidópteros y posterior purificación de los  productos de interés

PATRONES DE MOVIMIENTO EN ESPERMATOZOIDES DE ALPACA (Vicugna pacos): OBSERVACIONES ex situ.

The sperm hyperactivation is a step of the spermatozoa capacitation and consists ona change in the pattern of movements, the variation of the angle of the head, and thelongitude and width of the movement of the tail. The objective of the present study wasto determine the difference in movement patterns of spermatozoa and the frequency ofhyperactive spermatozoa in the oviduct according to the time after mating. The recoveryof the spermatozoa in five alpacas was done after 16, 20, 24, 28 and 32 hours post mating.Sperms were evaluated according to the pattern of movement. The frequency ofhyperactive spermatozoa increased until 28 hours post copula (71.4%).La hiperactivación espermática es un paso de la capacitación de los espermatozoides, consistente en un cambio en el patrón de movimiento, destacando la variación del ángulo de la cabeza, la longitud y amplitud del movimiento del flagelo. El objetivo del presente estudio fue determinar la diferencia entre los patrones de motilidad espermática, así como la frecuencia de espermatozoides hiperactivados en el oviducto, según el tiempo post cópula. La recuperación de los espermatozoides del oviducto de cinco alpacas se hizo a las 16, 20, 24, 28 y 32 horas post cópula. Los espermatozoides se evaluaron en base a su patrón de movimiento. El porcentaje de espermatozoides hiperactivados se incrementó hasta las 28 horas post cópula (71.4%)

Diagnostic efficacy of the ELISA test for the detection of deamidated anti-gliadin peptide antibodies in the diagnosis and monitoring of celiac disease

Volcano plot of differential gene expression in IZE_Suc vs IZE, ZE_5w vs IZE_Suc and SE_5w vs IZE. (DOCX 256 kb