Rapid detection of specific polyclonal and monoclonal antibodies against bovine leukemia virus

Llames, L.; Goyache, J.; Doménech, Ana; Ávila, Ana de; Suárez, G.; Gómez-Lucı́a, Esperanza

doi:10.1016/s0166-0934(99)00092-0

Cited by 11 publications

(7 citation statements)

References 23 publications

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“…These data conformed to other studies that assessed that dot blot assays are most sensitive and specific for the detection of low titers of antibodies directed against bacterial lipopolysaccharide (28) or viral antigens (20). The discrepancy between these two methods is related to the high nonspecific binding of samples in ELISA plates.…”

Section: Discussionsupporting

confidence: 86%

“…The discrepancy between these two methods is related to the high nonspecific binding of samples in ELISA plates. The replacement of the plastic binding matrix by a nitrocellulose membrane essentially solved this problem (20,28). In addition, ELISA had the disadvantage of being a test that required expensive equipment, making it difficult to adapt in the field (8).…”

Section: Discussionmentioning

confidence: 99%

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Dot Enzyme-Linked Immunosorbent Assay for More Reliable Staging of Patients with Human African Trypanosomiasis

et al. 2005

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Human African trypanosomiasis (HAT) or sleeping sickness is a disease characterized by a hemolymphatic stage 1 followed by a meningoencephalitic stage 2 which is fatal without specific treatment. Furthermore, due to the toxicity of drugs used to treat stage 2 (mainly melarsoprol) accurate staging is required. Actual criteria employed during field surveys are not sensitive enough for precise staging. Antineurofilament (anti-NF) and antigalactocerebrosides (anti-GalC) antibodies have been identified in cerebrospinal fluid (CSF) as potential markers of central nervous system (CNS) involvement. We describe a dot enzyme-linked immunosorbent assay (dot-ELISA) to detect anti-GalC and anti-NF antibodies and its value in staging. NF-and GalC-dotted nitrocellulose strips were first developed in our laboratory. They were then evaluated in Angola and Central African Republic on 140 CSF samples. Compared to our staging criteria (i.e., CSF cell count > 20 cells/l, CSF immunoglobulin M concentration > 100 mg/liter, and/or the presence of trypanosomes in the CSF), combined detection of both CSF anti-NF and CSF anti-GalC by dot-ELISA showed 83.2% sensitivity and 100.0% specificity. Dot-ELISA could be a useful test to diagnose CNS involvement in HAT in the less-equipped laboratories or in the field situation and to improve patient treatment.

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Section: Discussionsupporting

confidence: 86%

Section: Discussionmentioning

confidence: 99%

Dot Enzyme-Linked Immunosorbent Assay for More Reliable Staging of Patients with Human African Trypanosomiasis

et al. 2005

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“…Hybridoma supernatants (undiluted), GTD supernatants (1 : 100), ascitic fluid (1 : 200) or serum samples (1 : 100 in PBS‐T) (0.5 µl of each) were applied and incubated for 1 h at room temperature. After washing three times with PBS‐T, HRPO‐labelled second antibody [rabbit anti‐mouse polyvalent IgM and IgG (1 : 2000); and, goat anti‐bovine IgG (1 : 8000)] was added and incubated for 1 h. Reactions were revealed by immersion in substrate containing di‐amino‐benzidine ( Llames et al, 1999).…”

Section: Methodsmentioning

confidence: 99%

Production and Characterization of Monoclonal Antibodies against Bovine Leukaemia Virus using Various Crude Antigen Preparations: a Comparative Study

Llames

Gómez-Lucı́a

Doménech

et al. 2000

Journal of Veterinary Medicine, Series B

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A total of 59 monoclonal antibodies (mAbs) specific against the bovine leukaemia virus (BLV) using different antigen preparations was produced. The five antigen preparations for immunizing BALB/c mice were: live cells (CEL), sonicated and ultracentrifuged cells (SOC), cell lysates (LYS), semi-purified BLV (PV), and formalin-treated cells (FOR) from two cell lines permanently infected with BLV (FLK-BLV and BLV-bat2). These viral component presentations were selected to obtain mAbs against specific BLV proteins: located on the cell surface (FOR and CEL), in free virus particles (PV) and intracellular viral proteins (SOC and LYS). Two antigen preparations (SOC and LYS) were lethal to the mice following the intravenous and intrasplenic routes. Six fusions were performed in this study that rendered specific antibodies against BLV. The highest number of hybridomas was produced with SOC; however, the majority of the hybridomas produced (> 90%) were against cellular proteins. Even though immunization with PV gave the lowest number of hybridomas, the majority of them were specific against BLV. Based on the reactivity of the mAbs in Western blot (WB), we classified the mAbs into five groups, namely anti-gp51SU (39 mAbs), anti-gp30TM (six mAbs), anti-Pr72env (nine mAbs), anti-Pr66gag-pro (one mAb) and anti-Prgag (four mAbs). A very high percentage of the mAbs produced (48 of 59) reacted with gp51SU, suggesting that this is the most immunogenic and accessible BLV protein presented in the different antigen preparations. The majority of our mAbs recognized more than one band in WB, suggesting that, aside from reacting with mature proteins, the mAbs also recognized viral precursors.

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“…Methods for drug testing should be easier and more convenient. In Dot-ELISA, antigens binding NC membrane were stable for at least 5 months at different storage temperatures (À20 C, 4 C, and room temperature) (Llames et al 1999;Rai et al 2004). Based on this advantage, the BSA-conjugated drugs could be prepared long before the test is performed and drug test could proceed easily.…”

Section: Discussionmentioning

confidence: 99%

A 96-well Plate Based Dot-ELISA Array for Simultaneous Detection of Multi-Drugs

Wang¹,

Wei²,

Jin³

et al. 2009

Analytical Letters

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Drug abuse is rising day by day and becoming a big problem in countries around the globe. The existing drug testing methods could not effectively cope with the problems in the sensitivity and specificity of the diversified drugs. Dot-ELISA, as a complement of ELISA, has been widely used in many fields. In this study, we optimized and applied the Dot-ELISA technique as a maneuverable assay for rapid detection of multiple drug abuses via a kind of 96-well plates-based Dot-ELISA array. In total 92 positive urine samples were tested with high-throughput and all results were confirmed by commercial drug testing kits.

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Rapid detection of specific polyclonal and monoclonal antibodies against bovine leukemia virus

Cited by 11 publications

References 23 publications

Dot Enzyme-Linked Immunosorbent Assay for More Reliable Staging of Patients with Human African Trypanosomiasis

Dot Enzyme-Linked Immunosorbent Assay for More Reliable Staging of Patients with Human African Trypanosomiasis

Production and Characterization of Monoclonal Antibodies against Bovine Leukaemia Virus using Various Crude Antigen Preparations: a Comparative Study

A 96-well Plate Based Dot-ELISA Array for Simultaneous Detection of Multi-Drugs

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