scholarly journals The Potential of O’nyong-nyong Virus Strain SG650 Murine Monoclonal Antibodies for Detection of O’nyong-nyong and Chikungunya Viruses

2021 ◽  
Vol 3 (1) ◽  
pp. 97-114
Author(s):  
Albina Makio ◽  
Lillian Musila ◽  
Eddy Okoth Odari ◽  
Juliette Rose Ongus ◽  
Rosemary Sang
Keyword(s):  
Monoclonal Antibodies ◽  
Culture Fluid ◽  
Neutralization Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
High Morbidity ◽  
Diagnostic Potential ◽  
Potential Efficacy ◽  
Commercial Kits ◽  
Set Up ◽  
Murine Monoclonal Antibodies

O’nyong-nyong virus (ONNV) and Chikungunya virus (CHIKV) are antigenically related alphaviruses responsible for febrile illnesses common to the tropics and associated with relatively high morbidity and mortality. Murine monoclonal antibodies (mAbs) targeting alphaviruses like Chikungunya have been developed and used to make commercially available kits. However, few studies have been conducted to develop antibodies specific to ONNV and no commercial kits are available for use in endemic regions where outbreak potential is high. We demonstrate the potential of in-house generated monoclonal antibodies against ONNV to detect both ONNV and CHIKV. The objective of this study was to generate mAbs using hybridoma technology, characterize the developed mAbs, determine their specificity against selected alphaviruses and check their diagnostic potential using an indirect IgG enzyme-linked immunosorbent assay (ELISA) and focus neutralization assay (FRNT50). BALB/c mice were immunized with ONNV purified proteins from ONNV infectious culture fluid. After four rounds of booster injections, the mice were sacrificed, spleen cells harvested and fused with parental myeloma cells then cultured in selective media and the successful hybrid clones with antibody-producing ability purified to yield the desired mAbs. Five monoclonal antibodies targeting the ONNV E1 protein of isotypes IgG2a/kappa, IgG2b/kappa and IgM/kappa (P1B12, P1E9, P1G11, P1B4 and P1G6) demonstrated a potential to detect both ONNV and CHIKV isolates by indirect IgG ELISA but no potential for neutralization of the viruses by FRNT50. This study demonstrates the potential efficacy of in-house serological tools as an alternative in the absence of commercial assays in screening and diagnosis of ONN and CHIK viruses which are often co-circulating. It is our recommendation that this work may be pursued further to design and optimize ELISA assays, using the developed mAbs, for the detection of both ONN and CHIK viruses in the research laboratory set-up

scholarly journals Development of Murine Monoclonal Antibodies for the Immunohistochemical Diagnosis of Systemic Bovine Aspergillosis

1996 ◽  
Vol 8 (1) ◽  
pp. 68-75 ◽  
Author(s):  
H. E. Jensen ◽  
B. Aalbaek ◽  
P. Lind ◽  
H. V. Krogh ◽  
P. L. Frandsen
Keyword(s):  
Monoclonal Antibodies ◽  
Polyclonal Antibodies ◽  
Fungal Species ◽  
Cross Reactivity ◽  
Water Soluble ◽  
Enzyme Linked Immunosorbent Assay ◽  
Immunohistochemical Techniques ◽  
Immunohistochemical Diagnosis ◽  
Murine Monoclonal Antibodies

Murine monoclonal antibodies (MAbs) against water-soluble somatic antigens (WSSA) and the wall fraction (WF) from Aspergillus fumigatus were produced by fusion of splenocytes from immunized BALB/c mice with mouse myeloma X63-Ag 8.653 cells. The supernatants of in vitro cultured hybridomas were initially screened for reactivity with the WSSA and the WF from A. fumigatus and WSSA of other fungi in an enzyme-linked immunosorbent assay (ELISA). Supernatants reacting only with A. fumigatus antigens were subsequently screened for homologous and heterologous reactivity with immunohistochemical techniques using formalin-fixed, paraffin-embedded tissues from experimentally infected mice. Because of a high immunohistochemical reactivity with homologous fungi, 4 MAbs raised against A. fumigatus WSSA and WF were selected for a further evaluation of cross-reactivity (diagnostic specificity) in immunohistochemical and immunoblotting assays. In immunohistochemical assays, all MAbs raised against WSSA cross-reacted heavily with a number of other fungal species. All 4 MAbs (MAb-WF-AF-1-4) raised against the WF reacted strongly with hyphae of Aspergillus spp.; hyphae of Scedosporium apiospermum were also strongly labeled by MAb-WF-AF-3 and-4. The 2 specifically reacting MAbs (MAb-WF-AF-1 and-2) were of the IgM biotype and were precipitating, and in immunoblotting experiments both bound to a 106-kD antigen of the WF, whereas they did not bind to WSSA of A. fumigatus. One of the 2 aspergillosis-specific MAbs (MAb-WF-AF-1) was used to screen 145 mycotic lesions of cattle. The diagnoses on bovine lesions obtained by MAb-WF-AF-1 were compared with results based on reactivity with heterologously absorbed polyclonal antibodies and, for some lesions, to culture results. In the vast majority of lesions ( n = 133), the MAb-WF-AF-1 and the polyclonal anti-Aspergillus antibodies reacted in a similar pattern, i.e., positively in 41 aspergillosis lesions and negatively in 92 zygomycotic lesions. Hyphae in 3 of 12 lesions that were not stained by the polyclonal antibodies reacted with the specific MAb-WF-AF-1; i.e., aspergillosis was diagnosed. The characteristics of the 2 MAbs (MAb-WF-AF-1 and-2) raised against the WF of A. fumigatus in ELISA and immunoblotting and immunohistochemical assays justify their application for the in situ diagnosis of systemic aspergillosis of cattle.

MEASUREMENT OF PLASMINOGEN ACTIVATOR INHIBITOR 1 (PAI-1) IN PLASMA WITH AN ELISA BASED ON TWO MURINE MONOCLONAL ANTIBODIES

1987 ◽  
Author(s):  
P J Declerck ◽  
M C Alessi ◽  
M Verstreken ◽  
E K O Kruithof ◽  
I Juhan-Vague ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Pregnant Women ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
First Trimester ◽  
Enzyme Linked Immunosorbent Assay ◽  
Plasminogen Activator Inhibitor 1 ◽  
Murine Monoclonal Antibodies ◽  
Activator Inhibitor ◽  
Pai 1

An enzyme-linked immunosorbent assay (ELISA) for quantitation of plasminogen activator inhibitor 1 (PAI-1) was developed, based on two murine monoclonal antibodies (MA-7D4B7 and MA-7F5), raised against purified PAI-1 from HT-1080 fibrosarcoma cells, which react with non-overlapping epitopes. MA-7D4B7 was coated on microtiter plates and bound PAI-1 antigen was quantitated with MA-7F5 conjugated with horseradish peroxidase.In normal plasma, collected on citrate at pH 7.4, the PAI-1 level is 27 ± 16 ng/ml (mean ± SD, n=ll), with a corresponding value of 19 ± 11 ng/ml (n=12) in plasma collected on acid citrate pH 4.5, which inhibits the release of PAI-1 from platelets. The lower limit of the assay in plasma is 2 ng/ml; the intra-assay, inter-assay and inter-dilution coefficients of variation are 5.2%, 8.0% and 7.1% respectively.This ELISA was used to measure PAI-1 levels in plasma (collected on citrate, pH 7.4) of non-pregnant women and of women at different stages of pregnancy. A progressive increase is observed : before: 20±9 ng/ml, n=7; first trimester: 25 ± 12 ng/ml, n=5; second trimester: 40 ± 25 ng/ml, n=ll; third trimester: 98 ± 46 ng/ml, n=13. A correlation coefficient of 0.70 is found between the duration of pregnancy and the PAI-1 level.Preliminary data indicate that the PAI-1 antigen level is increased in several disease states, including myocardial infarction and deep vein thrombosis.Thus, this newly developed ELISA allows a direct measurement of the fast-acting inhibitor of plasminogen activator in plasma. Application of this assay to plasma of non-pregnant and pregnant women substantiates previous results obtained with the use of functional assays. In order to quantitate PAI-1 antigen circulating in plasma, blood should be collected under conditions that prevent platelet stimulation.

scholarly journals Detection of Human Papillomavirus Type 31-Neutralizing Antibodies from Naturally Infected Patients by an Assay Based on Intracellular Assembly of Luciferase-Expressing Pseudovirions

2007 ◽  
Vol 15 (1) ◽  
pp. 172-175 ◽  
Author(s):  
Maxime J. J. Fleury ◽  
Antoine Touzé ◽  
Silvia de Sanjosé ◽  
F. Xavier Bosch ◽  
Joellen Klaustermeiyer ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Human Papillomavirus ◽  
Immunosorbent Assay ◽  
Neutralizing Antibodies ◽  
Neutralization Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Highly Sensitive

ABSTRACT The aim of this study was to develop a highly sensitive human papillomavirus type 31 (HPV31) neutralization assay based on the production of pseudovirions carrying luciferase. Neutralizing antibodies against HPV31 were investigated in a set of HPV31 monoclonal antibodies and in women with evidence of HPV31 infection. Neutralizing antibodies were detected in 78% of subjects with a positive enzyme-linked immunosorbent assay.

scholarly journals Production of Specific Monoclonal Antibodies toAspergillus Species and Their Use in Immunohistochemical Identification of Aspergillosis

1999 ◽  
Vol 37 (4) ◽  
pp. 1221-1223 ◽  
Author(s):  
L. E. Fenelon ◽  
A. J. Hamilton ◽  
J. I. Figueroa ◽  
M. A. Bartholomew ◽  
M. H. Allen ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Immunoperoxidase Staining ◽  
Frozen Sections ◽  
Clinical Specimens ◽  
Murine Monoclonal Antibodies ◽  
Immunohistochemical Identification

Two anti-Aspergillus murine monoclonal antibodies (MAbs), designated 164G and 611F, have been produced; both specifically recognize cytoplasmic antigens of A. fumigatus, A. flavus, and A. niger by enzyme-linked immunosorbent assay. The MAbs can identify Aspergillus spp. both in frozen sections by immunofluorescence and in paraffin-embedded clinical specimens by immunofluorescence and immunoperoxidase staining.

scholarly journals Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus Infections

Pathogens ◽  
2019 ◽  
Vol 8 (3) ◽  
pp. 140
Author(s):  
Xueming Chen ◽  
Tongtong Li ◽  
Xiaodan Chen ◽  
Chenxi Li ◽  
Weiwei Lin ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Core Protein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dependent Manner ◽  
Serum Samples ◽  
Independent Manner ◽  
Competitive Binding Assay ◽  
Diagnostic Potential ◽  
Peptide Phage Display

Although σA is an important major core protein of duck reovirus (DRV), the B-cell epitopes of this protein remain unknown to reseacrhers. Six monoclonal antibodies (MAbs) (1A7, 3F4, 5D2, 4E2, 3C7, and 2B7) were developed by using prokaryotic-expressed recombinant His-σA protein. Five of six MAbs (1A7, 3F4, 4E2, 3C7, and 2B7) reacted with His-σA protein in a conformation-independent manner, while 5D2 reacted with σA in a conformation-dependent manner. Immunofluorescence assays showed that the MAbs could specifically bind to DRV infected BHK-21 cells. The MAbs were delineated as three groups by a competitive binding assay. By using 12-mer peptide phage display and mutagenesis, MAb 4E2 was identified to recognize minimal epitope 56EAPYPG61 and MAb 1A7 recognize 341WVV/MAGLI/V347, residues 341V/M and 347I/V are replaceable. Dot blotting and sequence analysis confirmed that EAPYPG and WVV/MAGLI/V are cross-reactive epitopes in both DRV and avian reovirus (ARV). An enzyme-linked immunosorbent assay (ELISA) based on two expressed EAPYPG and WVVAGLI as antigen demonstrated its diagnostic potential by specific reacting with serum samples from DRV- or ARV-infected birds. Based on these observations, an epitope-based ELISA could be potentially used for DRV or ARV surveillance. These findings provide insights into the organization of epitopes on σA protein that might be valuable for the development of epitope-based serological diagnostic tests for DRV and ARV infection.

scholarly journals New Monoclonal Antibodies for a Selective Detection of Membrane-Associated and Soluble Forms of Carbonic Anhydrase IX in Human Cell Lines and Biological Samples

Biomolecules ◽  
2019 ◽  
Vol 9 (8) ◽  
pp. 304 ◽  
Author(s):  
Dovile Stravinskiene ◽  
Aiste Imbrasaite ◽  
Vilma Petrikaite ◽  
Daumantas Matulis ◽  
Jurgita Matuliene ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Carbonic Anhydrase ◽  
Tumor Cells ◽  
Cell Lines ◽  
Carbonic Anhydrase Ix ◽  
Soluble Form ◽  
Enzyme Linked Immunosorbent Assay ◽  
Tumor Biomarker ◽  
Ca Ix ◽  
Diagnostic Potential

Monoclonal antibodies (MAbs) selectively targeting tumor-associated antigens such as carbonic anhydrase IX (CA IX) can significantly contribute to research, diagnostics, and treatment of CA IX-related cancers. CA IX is overexpressed in numerous hypoxic cancers where it promotes tumor progression. Therefore, it is considered as a promising tumor biomarker. A novel collection of MAbs against recombinant CA IX was developed and evaluated in different immunoassays for studying CA IX expression. The reactivity of MAbs with native cell surface protein was confirmed by flow cytometry and the presence of hypoxia-inducible CA IX was investigated in several human cancer cell lines. In addition, the applicability of MAbs for visualization of CA IX-positive tumor cells by immunofluorescence microscopy was demonstrated. MAb H7 was identified as the most promising MAb for different immunoassays. It recognized a linear epitope covering CA IX sequence of 12 amino acid residues 55-GEDDPLGEEDLP-66 within the proteoglycan domain. The MAb H7 was the only one of the collection to immunoprecipitate CA IX protein from cell lysates and detect the denatured CA IX with near-infrared fluorescence Western blot. It was also employed in sandwich enzyme-linked immunosorbent assay to detect a soluble form of CA IX in growth medium of tumor cells and blood plasma samples. The diagnostic potential of the MAb H7 was confirmed on formalin-fixed and paraffin-embedded tissue specimen of cervical carcinoma in situ by immunohistochemistry. The generated MAbs, in particularly clone H7, have great potential in diagnostics and research of CA IX-related cancers.

Diagnostic Potential of Monoclonal Antibodies to Adenovirus

2021 ◽  
Vol 37 (2) ◽  
pp. 48-53
Author(s):  
I.V. Amosova ◽  
M.P. Grudinin
Keyword(s):  
Monoclonal Antibodies ◽  
Sandwich Elisa ◽  
Human Adenovirus ◽  
Clinical Samples ◽  
Enzyme Linked Immunosorbent Assay ◽  
Hexon Protein ◽  
Diagnostic Potential ◽  
Immunological Tests ◽  
Cell Enzyme ◽  
Cell Elisa

The diagnostic potential of 4B7 and 6B12 monoclonal antibodies (mAbs) against human adenovirus hexon protein has been studied in various immunological tests, namely: in-cell enzyme-linked immunosorbent assay (cell-ELISA), sandwich ELISA, and immunofluorescent assay (IFA). It was shown that the sensitivity of cell-ELISA, sandwich ELISA and IFA was 96%, 86% and 84%, respectively as compared to PCR. Thus, 4B7 and 6B12 mAbs are promising immunoreagents for the construction of various diagnostic kits to use in laboratory practice for adenovirus detection in clinical samples. monoclonal antibodies, human adenovirus, adenovirus infection, diagnosis

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