scholarly journals Application of in situ PCR for the Detection of Bovine Leukaemia Virus (BLV) Infection in Dendritic Cell cultures

2014 ◽  
Vol 58 (3) ◽  
pp. 347-352 ◽  
Author(s):  
Ewelina Iwan ◽  
Maria Szczotka ◽  
Jacek Kuźmak
Keyword(s):  
Cell Cultures ◽  
Nested Pcr ◽  
Leukemia Virus ◽  
Positive Control ◽  
Adherent Cell ◽  
Bovine Leukaemia Virus ◽  
Leukaemia Virus ◽  
Pcr Method

Abstract The aim of the study was to develop an in situ PCR (IS-PCR) method for detection of bovine leukemia virus (BLV) in cell cultures. Samples from five BLV positive and five BLV negative cows were collected and dendritic cells (DCs) from blood, bone marrow, spleen, and lymph node were cultured. Cultures prepared from healthy animals were infected with BLV. After two weeks, the cells were tested by nested PCR and IS-PCR for the presence of proviral DNA. As a positive control adherent cell line permanently infected with BLV was used. BLV was successfully detected by IS-PCR in DCs from naturally infected cattle and DCs infected in vitro. In control, non-infected DCs, the results of the reaction were negative. The results of provirus detection by IS-PCR were similar with these performed with nested PCR. Additionally, IS-PCR provides many advantages, like specific localisation of infection and smaller number of cells needed as template for PCR.

Hepatoma Cell Cultures as In Vitro Models for the Hepatotoxicity of Xenobiotics

1988 ◽  
Vol 16 (1) ◽  
pp. 32-37
Author(s):  
Margherita Ferro ◽  
Anna Maria Bassi ◽  
Giorgio Nanni
Keyword(s):  
Cell Lines ◽  
Cell Cultures ◽  
Membrane Lipids ◽  
Hepatoma Cell ◽  
Positive Control ◽  
In Vitro Models ◽  
Low Concentrations ◽  
Formation Efficiency ◽  
Dose Dependent

Two hepatoma cell cultures were examined as in vitro models to be used in genotoxicity and cytotoxicity tests without the addition of bioactivating enzymes. The MH1C1, and HTC hepatoma lines were used in this study to establish their sensitivity to a number of xenobiotics, namely, cyclophosphamide (CP), the classical positive control in bioactivation tests; benzaldehyde (BA), a short-chain aldehyde; and 4-hydroxynonenal (HNE), a major toxic end-product of the peroxidative degradation of cell membrane lipids. As a first approach, we compared the following cytotoxicity tests: release of lactate dehydrogenase (LDH), and colony formation efficiency (CF). Colony-forming cells were exposed to the drugs according to different procedures, before or after the anchorage phase. The leakage of LDH into the medium following exposure of both cell lines to HNE, CP and BA for up to 24 hours was found not to be a good index of cytotoxicity. A better indicator of cytotoxicity was CF, as evaluated by exposure of the cells 24 hours after seeding. The effects were detectable at very low concentrations, corresponding to 10, 90 and 100μM for HNE, CP and BA, respectively. The impairment of CF efficiency was dose-dependent and time-dependent, and several differences between the two cell lines were observed.

Infectivity of bovine leukaemia virus infected cattle: An ELISA for detecting antigens expressed in in vitro cultured lymphocytes

1992 ◽  
Vol 30 (2-3) ◽  
pp. 137-150 ◽  
Author(s):  
J.A. Cowley ◽  
J.B. Molloy ◽  
C.K. Dimmock ◽  
P.J. Walker ◽  
A.G. Bruyeres ◽  
...  
Keyword(s):  
Infected Cattle ◽  
Bovine Leukaemia Virus ◽  
Leukaemia Virus

scholarly journals Electrical impedance measurement system to assess in-situ experiments on epithelia under ELF magnetic fields exposure

2021 ◽  
Vol 19 (3) ◽  
pp. 217-226
Author(s):  
G. Domínguez ◽  
E. Cardiel ◽  
J.L Reyes ◽  
E. Sánchez ◽  
P.R. Hernández
Keyword(s):  
Impedance Spectroscopy ◽  
Magnetic Fields ◽  
Cell Cultures ◽  
Electrical Impedance ◽  
Controlled Environment ◽  
Spectroscopy Technique ◽  
In Situ Experiments ◽  
Impedance Spectroscopy Technique

Purpose: The development of an electric impedance meter based on the impedance spectroscopy technique, for in vitro and in situ experimentation, with cellular epithelia submitted to extremely low frequency magnetic fields in a controlled environment. Unlike other reported systems, a strength of the one presented here is that it avoids the influence of external factors on the experiment. Materials and methods: The designed system employs the electrical impedance values obtained by the impedance spectroscopy technique to determine the parameters of the simple equivalent electrical model of a cellular monolayer. The Madin-Darby Canine Kidney (MDCK) cell cultures were used as subjects of study in the experimental protocol. Results: The validation was carried out by comparing the transepithelial electrical impedance data of the cell cultures obtained with the developed system and those of the Cellzscope® commercial system used as the standard. Non-significant differences were obtained. Conclusion: It was confirmed that the developed system provides reliable values of transepithelial electrical impedance to experiment with cell cultures and take advantage of the controlled environment to reduce the effects of experimental management.

scholarly journals Sex determining of cat embryo and some feline species

Zygote ◽  
2008 ◽  
Vol 16 (2) ◽  
pp. 169-177 ◽  
Author(s):  
Francesca Ciani ◽  
Natascia Cocchia ◽  
Maria Rizzo ◽  
Patrizia Ponzio ◽  
Gennaro Tortora ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Positive Control ◽  
Domestic Cat ◽  
Preimplantation Embryos ◽  
Pcr Analysis ◽  
Domestic Cats ◽  
Hair Samples ◽  
Wild Felids

SummarySex identification in mammalian preimplantation embryos is a technique that is used currently for development of the embryo transfer industry for zootechnical animals and is, therefore, a resource for biodiversity preservation. The aim of the present study was to establish a rapid and reliable method for the sexing of preimplantation embryos in domestic cats. Here we describe the use of nested PCR identify Y chromosome-linked markers when starting from small amounts of DNA and test the method for the purpose of sexing different species of wild felids. To evaluate the efficiency of the primers, PCR analysis were performed first in blood samples of sex-known domestic cats. Cat embryos were produced both in vitro and in vivo and the blastocysts were biopsied. A Magnetic Resin System was used to capture a consistent amount of DNA from embryo biopsy and wild felid hairs. The results from nested PCR applied on cat blood that corresponded to the phenotypical sex. Nested PCR was also applied to 37 embryo biopsies and the final result was: 21 males and 16 females. Furthermore, β-actin was amplified in each sample, as a positive control for DNA presence. Subsequently, nested PCR was performed on blood and hair samples from some wild felines and again the genotyping results and phenotype sex corresponded. The data show that this method is a rapid and repeatable option for sex determination in domestic cat embryos and some wild felids and that a small amount of cells is sufficient to obtain a reliable result. This technique, therefore, affords investigators a new approach that they can insert in the safeguard programmes of felida biodiversity.

Particokinetics and in vitro dose of high aspect ratio nanoparticles

Nanoscale ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 5209-5214 ◽  
Author(s):  
Seth Richard Price ◽  
Calum Kinnear ◽  
Sandor Balog
Keyword(s):  
Aspect Ratio ◽  
Cell Cultures ◽  
High Aspect Ratio ◽  
Adherent Cell

Adapting computational particokinetic models to address the dosage of high-aspect ratio nanomaterials for in vitro nanoparticle toxicology assays involving submerged adherent cell cultures.

scholarly journals Comparison of agar gel immunodiffusion test, enzyme-linked immunosorbent assay and PCR in diagnostics of enzootic bovine leukosis

2005 ◽  
Vol 59 (3-4) ◽  
pp. 363-370
Author(s):  
Tadej Malovrh ◽  
M. Pate ◽  
M. Ocepek ◽  
B. Krt
Keyword(s):  
Antibody Response ◽  
Immunosorbent Assay ◽  
Large Scale ◽  
Serological Test ◽  
Enzyme Linked Immunosorbent Assay ◽  
Agar Gel ◽  
Bovine Leukaemia Virus ◽  
Leukaemia Virus ◽  
Elisa Kit ◽  
Pcr Method

Bovine leukaemia virus (BLV) is a retrovirus that induces a chronic infection in cattle. Once infected, cattle remain virus carriers for life and start to show an antibody response within a few weeks after infection. Eradication and control of the disease are based on early diagnostics and segregation of the carriers. The choice of a diagnostic method depends on the eradication programme, money resources and characteristics of the herd to be analysed. The agar gel immunodiffusion (AGID) test has been the serological test of choice for routine diagnosis of serum samples. Nevertheless, in more recent years, the enzyme-linked immunosorbent assay (ELISA) has replaced the AGID for large scale testing. For this purpose, commercially available BLV-ELISA kits were compared to the AGID and to the polymerase chain reaction (PCR) method performed with two sets of primers, amplifying env region. The ELISA kit based on the p24 core protein was found to be less specific and served as a screening test. The ELISA kit based on the envelope glycoprotein (gpSI) served as a verification test and gave a good correlation with the AGID test and PCR method. However, ELISA showed a higher sensitivity than AGID. The p24 based ELiSA was useful for screening a large number of samples, whereas gp51 based ELISA, AGID and PCR were more important for detecting the antibody response against the individual BLV-proteins and therefore for verification of the infection with BLV.

Replication of type C Virus Particles in Burkitt Lymphoma Cells in Vitro Treated with Feline or Murine Leukemia Virus

1970 ◽  
Vol 28 ◽  
pp. 164-165
Author(s):  
K. Maruyama ◽  
T. Fujimoto ◽  
G. R. Swearingen ◽  
L. Dmochowski
Keyword(s):  
Cell Cultures ◽  
Burkitt Lymphoma ◽  
Murine Leukemia Virus ◽  
Human Cancer ◽  
Leukemia Virus ◽  
Culture Fluid ◽  
Type C ◽  
Human Embryo Kidney ◽  
Murine Leukemia

In the course of studies on possible interaction between animal leukemia viruses and a hypothetical human cancer virus(es), a culture line (P3) derived from Burkitt lymphoma and containing herpestype virus was inoculated 10 times with feline or murine leukemia virus. Filtrates (0.45 µ Millipore) of tissue culture fluid from feline lymphoma cell cultures containing type C particles (supplied by Dr. C. G. Rickard) and from human embryo kidney cell cultures infected with Rauscher leukemia virus(supplied by Dr. B. S. Wright) were used as source of virus. Electron micro scopy was carried out on cells from cultures before inoculation, the 4-day-old culture after 10 inoculations, and subcultures at 2nd (30 days), 3rd (40 days), 12th(85 or 95 days) passages after the last inoculation.

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