Development and validation of an ELISA for quantitation of bovine viral diarrhea virus antigen in the critical stages of vaccine production.

Bovine Viral Diarrhea Virus (BVDV) is the causative agent of a worldwide disease. The virus infects bovines of all ages, causing reproductive problems and contaminating biological products of high commercial value. The large-scale production of BVDV vaccines presents the challenge of processing antigenic proteins that are highly susceptible to the processing environment. Potency testing requires the immunization of cattle in order to determine the neutralizing antibodies titers induced by the vaccine. An alternative to the in vivo test is an in vitro measurement of key viral antigens. This paper describes the development and validation of a sandwich-type indirect ELISA that is able to detect and quantify BVDV E2 glycoprotein in live and inactivated BVDV. Validation parameters such as repeatability, intermediate precision, and reproducibility indicated that the developed ELISA constitutes an advanced tool for evaluating the BVDV antigen throughout manufacturing and vaccine release testing.

Truncated E2 of bovine viral diarrhea virus (BVDV) expressed in Drosophila melanogaster cells: a candidate antigen for a BVDV ELISA.

A simple and reliable indirect enzyme-linked immunosorbent assay for detection of antibodies directed against a major bovine viral diarrhea virus (BVDV) immunogen, the E2 glycoprotein (tE2-ELISA), has been developed using the recombinant C-terminal truncated E2 glycoprotein (tE2) expressed in a Drosophila melanogaster system. This strategy demonstrated that tE2 is secreted efficiently in the supernatant, no purification steps are necessary, it is easy to produce and carries out the post translational modifications necessary to preserve its native conformation. Preliminary analysis of 183 cattle serum samples using tE2-ELISA showed a 98% specificity and a 100% sensitivity compared with the standard homologous BVDV virus neutralization test. The results also showed that the tE2 is immunoreactive because the conformation and antigenicity of the original E2 are maintained to a large extent. To our knowledge this is the first study report of the recombinant tE2 of BVDV expressed in D. melanogaster system as an antigen for ELISA.

Evidence that the cannabinoid CB1 receptor is a 2-arachidonoylglycerol receptor. Structure-activity relationship of 2-arachidonoylglycerol, ether-linked analogues, and related compounds.

An endogenous cannabimimetic molecule, 2-arachidonoylglycerol, induces a rapid, transient increase in intracellular free Ca2+ concentrations in NG108-15 cells through a cannabinoid CB1 receptor-dependent mechanism. We examined the activities of 24 relevant compounds (2-arachidonoylglycerol, its structural analogues, and several synthetic cannabinoids). We found that 2-arachidonoylglycerol is the most potent compound examined so far: its activity was detectable from as low as 0.3 nM, and the maximal response induced by 2-arachidonoylglycerol exceeded the responses induced by others. Activities of HU-210 and CP55940, potent cannabinoid receptor agonists, were also detectable from as low as 0.3 nM, whereas the maximal responses induced by these compounds were low compared with 2-arachidonoylglycerol. Anandamide was also found to act as a partial agonist in this assay system. We confirmed that free arachidonic acid failed to elicit a response. Furthermore, we found that a metabolically stable ether-linked analogue of 2-arachidonoylglycerol possesses appreciable agonistic activity, although its activity was apparently lower than that of 2-arachidonoylglycerol. We also confirmed that pretreating cells with various cannabinoid receptor agonists nullified the response induced by 2-arachidonoylglycerol, whereas pretreating cells with other neurotransmitters or neuromodulators did not affect the response. These results strongly suggested that the cannabinoid CB1 receptor is originally a 2-arachidonoylglycerol receptor, and 2-arachidonoylglycerol is the intrinsic physiological ligand for the cannabinoid CB1 receptor.

Myocardial perfusion with [11C]methyl triphenyl phosphonium: measurements of the extraction fraction and myocardial uptake.

The present study describes extraction fraction and uptake measurements of the [11C]methyl triphenyl phosphonium (11C-MTP), a promising positron emission tomography (PET) agent for cardiac imaging. PET imaging was performed in mongrel dogs. Under physiological flow conditions 11C-MTP uptake reached a maximum within the first 10 minutes after injection and remained constant during the entire observation period of 80 minutes. Over the same time period, the heart/blood ratio was 46-106:1, and the heart/lung ratio 14:1. Following permanent occlusion of the left anterior descending coronary artery, 11C-MTP uptake in the normally perfused myocardium also reached a maximum at 10 minutes after injection, whereas in the infarcted area there was no significant accumulation of 11C-MTP. For a time period of 80 minutes the noninfarcted/infarcted myocardium ratio was 12:1. Extraction was measured in anesthetized dogs with a double isotope method using 99mTc-HSA as the reference tracer. The extraction fraction was 91% at a flow of 69 mL/min/100g. As flow increased to five-fold (342 mL/min/100g) following administration of adenosine, extraction fell to 61%. Following coronary artery occlusion, the 11C-MTP content in the myocardium was highly correlated (r = 0.93, p < 0.01; y = 10.46 + 0.92x) with the microsphere determined regional myocardial blood flow.

Large-scale use of liquid-phase blocking sandwich ELISA for the evaluation of protective immunity against aphthovirus in cattle vaccinated with oil-adjuvanted vaccines in Argentina.

Specific serum activity levels against four reference strains of foot-and-mouth disease virus (FMDV) were evaluated from 1634 animals vaccinated with commercial quadrivalent oil vaccines and from 746 unvaccinated, naive animals, using the liquid-phase blocking sandwich ELISA (lpELISA) test. Cows from the FMDV-free area of Argentina were tested for the absence of specific FMDV antibodies (sp FMDV Abs) and those showing lpELISA titres < 1.0 were grouped in lots of 16 animals. They were vaccinated and challenged at 90 days postvaccination (DPV) with one of four virus strains used for vaccine production and control (prototype strains). Serum samples from vaccinated and control cattle were collected 60 and 90 DPV and the level of sp FMDV Abs was determined by lpELISA. Animals were examined for clinical signs of disease. Results show that serum lpELISA titre levels directly correlate with the percentage of protected animals. It was seen that 100, 98, 93 and 87% of the vaccinated cattle with antibody titre levels > or = 2.1 were protected against challenge with serotypes C85, A87,01 Cas and A79, respectively. Evidence is also presented of seroconversion in a sample of 3-5-month-old calves vaccinated in the field, showing lpELISA titres compatible with protection against the four vaccine viruses as long as 150 DPV. Results reported in this paper strongly support the use of the lpELISA test for a rapid and reliable evaluation of the efficacy of FMDV commercial vaccines as well as for the assessment of the immunological status of cattle in FMDV-free and enzootic regions of South America.

[Survey of histopathological diagnostic services in the Department of Oral Pathology, School of Dentistry, Higashi-Nippon-Gakuen University, 1979-1989].

This study is based on an analysis of oral biopsy specimens that were diagnosed histopathologically in the Department of Oral Pathology, School of Dentistry, HIGASHI-NIPPON-GAKUEN UNIVERSITY from 1979-1989. 1. The total number of specimens during the ten years were 503 (dental hospital: 363, other personal dental clinics: 140), it corresponding to 440 cases (patients). 2. There were 224 men, 210 women, and 6 unknown. 3. The most common cases were radicular cysts 11.1% (49 cases), the second dental granuloma 9.5% (42 cases), followed by fibroma (fibrous polyp) 7.7% (34 cases). 4. The largest group of all specimens were cysts (35.7%), the second inflammatory lesions (28.0%), followed by tumors (26.6%). 5. About 80% of oral tumors were benign and about 20% were malignant. Of 24 cases with malignant tumors, the majority were squamous cell carcinoma (20 cases, 87%), followed by malignant melanoma (2 cases, 8.7%), verrucous carcinoma (1 case, 4.3%), and clear cell carcinoma (1 case, 4.3%). 6. About 13% of all oral tumors were odontogenic tumors, and the most common were ameloblastoma (8 cases, 53.3%), followed by odontoma (5 cases, 33.3%), and cementifying fibroma (2 cases, 13.3%). 7. The most common cases of specimens from other dental clinics were radicular cysts (23 cases, 16.4%), the second fibroma (fibrous polyp) (17 cases, 12.1%), and followed by dental granuloma (15 cases, 10.1%).

Resistencia de Crotalus durissus terificus y Bothrops neuwiedii a la neurotoxidad de cantidades masivas de veneno crotalico. / Resistance of Crotalus durissus terrificus and Bothrops neuwiedii to neurotoxicity of massive quantities of Crotalid vevenom

Especimenes de Crotalus durissus terrificus y de Bothrops neuwiedii toleran cantidades de veneno crotalico equivalentes a 5 x 10 elevado a menos 4 de su peso.La tolerancia observada puede explicarse por la presencia en el suero de ambas especies de componente (s) capaces de inactivar crotoxina, la neurotoxina mayor del veneno crotalico. La potencia antitoxica del suero antitoxica del suero de C. d. terrificus frente al veneno crotalico es similar a la del suero antiofidico monovalente obtenido de caballo hiperinmune, mientras que la potencia antitoxica del suero de B. neuwiedii es de alredor del 20% del de este ultimo. Los sueros de ambas especies en concentraciones adequadas son capaces de de proteger al raton frente a 4 LD 50 de veneno crotalico. La ausencia de lineas de precipitacion en ensayo de doble difusion de suero frente a veneno sugiere que los factores antitoxicos de ambas especies no son inmunoglobulinas. Es probable que la crotoxina sea neutralizada por la formacion de complexos inactivos con componentes especificos del suero. La resistencia al veveno no es reciproca, ya que especimenes de C.d. terrificus mueren luego de la inyeccion de cantidades de veneno de B.neuwiedii que son perfectamente toleradas por la especie dadora

Comunicacao sobre fluorose. / Fluorinese

Esta comunicacao refere-se a identificacao do risco da fluorose, tendo-se verificado a aquisicao desta por trabalhadores de industrias de fertilizantes em Cubatao. O risco foi constatado com base na analise de fluoretos urinarios. Recomenda-se que os colegas se mantenham atentos ao problema em outras regioes, especialmente em fabricas ou mineracoes, onde possa haver o risco