Chemiluminescence: sensitive detection technology for reporter gene assays

A series of enzyme-activated chemiluminescence-based assays of reporter gene expression, useful in many biomedical applications, has been developed. The chemiluminescence detection systems for beta-galactosidase, beta-glucuronidase (GUS), and secreted placental alkaline phosphatase (SEAP) reporter enzymes are all based on use of 1,2-dioxetane substrates. This detection technology also permits the combined luminescence detection of two different reporter enzymes in the same tube, e.g., a dual assay for beta-galactosidase and luciferase. The sensitivity of these chemiluminescence assays is several orders of magnitude greater than that of conventional colorimetric or fluorometric detection methods; e.g., the detection limit for beta-galactosidase by the chemiluminescence assay is 8 fg and by a fluorometric assay is 2 pg. Furthermore, chemiluminescence enables detection of beta-galactosidase, GUS, and SEAP enzyme concentrations over a dynamic range of more than five to six orders in magnitude. These assays offer highly sensitive, quantitative, rapid, nonisotopic detection of reporter enzymes that are widely used in both mammalian cells and plant cells.     

Seroprevalence to bovine immunodeficiency virus and lack of association with leukocyte counts in Italian dairy cattle

We report herein on the first serological detection of antibodies to bovine immunodeficiency virus (BIV) in Italy. According to criteria of a stratified-random sampling of dairy cattle reared in the Parma area (a province in the Po Valley, Northern Italy), sera from 3166 cows belonging to 272 herds were collected. In addition, sera of 138 bulls from eight artificial-insemination (AI) centres were sampled. Seventy-eight cows (2.5%) from 16 herds (5.8%) and seven bulls (5.1%) from two AI centres were positive for BIV-R29 antibodies in the IFA-test. IFA-positive sera assayed by Western blot had reaction to different viral proteins: 81 out of 85 sera showed antibody to p26 (considered the BIV major internal core protein); four sera reacted to other viral proteins but not to p26. Peripheral blood leukocytes of 60 seropositive and 60 seronegative animals, belonging to eight BIV-infected herds, were enumerated to assess any effect of BIV infection on white-blood cells. No significant differences were detected between the two groups. These data indicate that BIV infection is present in Italian dairy cattle--but the role of BIV in inducing disease remains unclear.     

Neurotrophin receptor expression is induced in a subpopulation of trigeminal neurons that label by retrograde transport of NGF or fluoro-gold following tooth injury

Tissue responses to injury are regulated by neurotrophins and neurotrophin receptor levels and can involve both retrograde and paracrine/autocrine trophic signaling. To determine how neurotrophins may contribute to the injury response, the timing and the extent of the up-regulation of neurotrophins and their receptors was examined in a model system which is particularly well suited for the analysis of trophic signaling pathways in response to injury. Injury to the occlusal surfaces of rat molar cusps induces a localized increase in nerve growth factor (NGF) expression in the dental pulp within 4-6 h. Radiolabeled NGF was transported in a receptor-mediated fashion from the teeth to a subset of neurons in the trigeminal ganglion within 15 h, indicating that these neurons possess NGF receptors (trk A and/or p75NTR). To test for NGF responses in the tooth sensory afferent neurons, levels of expression of neurotrophins and their receptors were examined by in situ hybridization in the trigeminal ganglion at 0, 4, 12, 20, 28 and 52 h post-injury. Within the maxillary division of the trigeminal ganglion, trk A expression was elevated at 4 h post-injury, with a maximum increase (2-fold) after 52 h. p75NTR was increased by 28 h post-injury and was increased 1.35-fold by 52 h. BDNF mRNA was increased 12 h after injury (1.8-fold), and 2.5-3-fold at 52 h post-injury. The trk B expression was increased only late after injury (28 and 52 h). To determine the receptor/neurotrophin phenotype of trigeminal neurons with projections to the molar teeth, these neurons were double-labeled with the retrograde tracer fluoro-gold and probes for either BDNF or trk B. The results show that tooth-innervating trigeminal neurons express BDNF, but not trk B. The timing of mRNA expression after injury and the phenotype of identified trigeminal neurons suggests a complex signaling cascade in which NGF at the injury site regulates NGF receptor expression at the levels of the cell body as well as increases in BDNF expression. Upregulated BDNF may act in a paracrine fashion on neighboring trigeminal cells expressing trk B. This signaling cascade may be a common feature of the response to mild peripheral inflammatory injuries within nociceptive pathways.     

Acute Q fever hepatitis in Taiwan

We report a case of a chronic hepatitis B carrier with an episode of acute hepatitis. The patient presented with a headache, arthralgias, jaundice and fever. While the laboratory tests mimicked chronic hepatitis B with an acute exacerbation, lipogranulomatous changes seen in the liver biopsy strongly suggested the presence of Q fever. Serology testing for Coxiella burneti proved positive and the patient responded to tetracycline therapy. While previously unreported in Taiwan, Q fever is important to consider in an atypical hepatitis presentation because it is a treatable condition.     

Studies on the molybdenum application to soybean

Treatment of inoculated soybean seeds with 2% ammonium molybdate solution brought about a significant increase in the yield and the N content of the plants. This was demonstrated in pot and field trials. It was also found that the beneficial effect of the Mo treatment was due to an increase in N fixation