scholarly journals Paracoccidioides HSP90 can be found in the cell surface and is a target for antibodies with therapeutic potential

2020 ◽  
Author(s):  
Ágata Nogueira D’Áurea Moura ◽  
Diane Sthefany Lima de Oliveira ◽  
Verenice Paredes ◽  
Letícia Barboza Rocha ◽  
Arturo Casadevall ◽  
...  
Keyword(s):  
Latin America ◽  
Monoclonal Antibodies ◽  
Cell Surface ◽  
Therapeutic Potential ◽  
Polyclonal Antibodies ◽  
Paracoccidioides Brasiliensis ◽  
Severe Disease ◽  
Rural Workers ◽  
Drug Candidates ◽  
Yeast Cell Surface

AbstractParacoccidioidomycosis (PCM) is one of the most frequent systemic mycoses in Latin America. It affects mainly male rural workers in impoverished regions, and the therapy can last up to two years or use drugs that are very toxic. Given the need for novel safe and effective approaches to treat PCM, we have been developing monoclonal antibodies (mAbs) that could be used not only to block specific fungal targets, but also modulate the host’s antifungal immunity. In this work we show the generation of and promising results with a mAb against HSP90, a molecular chaperone that is an important virulence factor in fungi. Using recombinant Paracoccidioides lutzii (Pb01) and P. brasiliensis (Pb18) HSP90 proteins produced in E. coli, we immunized mice and generated polyclonal antibodies and an IgG1 hybridoma mAb. The proteins were very immunogenic and both the polyclonal serum and mAb were used in immunofluorescence experiments, which showed binding of antibodies to the yeast cell surface. The mAb successfully opsonized P. lutzii and P. brasiliensis cells in co-incubations with J774.16 macrophage-like cells. Our results suggest that this mAb could serve as the basis for new immunotherapy regimens for PCM.Author summaryParacoccidioidomycosis (PCM) is a severe disease caused by fungi, common in Latin America. It is treatable, but some of the drugs that are available are very toxic or not very effective, and the treatment can take as long as two years to clear the infection. To address the need for improved therapeutic alternatives, we have been developing drug candidates based on antibody technologies against Paracoccidioides brasiliensis and P. lutzii, which cause PCM. In this work, we produced monoclonal antibodies (mAbs) that bind to the fungal protein HSP90, which is essential for fungal cells to survive. One mAb, 4D11, recognized the HSP90 target on the surface of fungal cells. These antibody-covered cells were ingested more efficiently by immune cells called macrophages, suggesting they could improve the host resistance to infection by Paracoccidioides. Future improvements on these antibodies could thus lead to more effective and safer PCM treatments.

scholarly journals Paracoccidioides HSP90 Can Be Found in the Cell Surface and Is a Target for Antibodies with Therapeutic Potential

2020 ◽  
Vol 6 (4) ◽  
pp. 193
Author(s):  
Ágata Moura ◽  
Diane Oliveira ◽  
Verenice Paredes ◽  
Letícia Rocha ◽  
Fabiana Oliveira ◽  
...  
Keyword(s):  
Cell Surface ◽  
Molecular Chaperone ◽  
Therapeutic Potential ◽  
Polyclonal Antibodies ◽  
E Coli ◽  
Rural Workers ◽  
Yeast Cell Surface ◽  
Important Virulence Factor ◽  
Hsp 90 ◽  
Polyclonal Serum

Paracoccidioidomycosis (PCM) is one of the most frequent systemic mycoses in Latin America. It affects mainly male rural workers in impoverished regions, and the therapy can last up to two years or use drugs that are very toxic. Given the need for novel safe and effective approaches to treat PCM, we have been developing monoclonal antibodies (mAbs) that could be used not only to block specific fungal targets, but also modulate the host’s antifungal immunity. In this work we show the generation of and promising results with an mAb against Heat Shock Protein (HSP)90, a molecular chaperone that is an important virulence factor in fungi. Using recombinant Paracoccidioides lutzii (Pb01) and P. brasiliensis (Pb18) HSP90 proteins produced in E. coli, we immunized mice and generated polyclonal antibodies and an IgG1 hybridoma mAb. The proteins were very immunogenic and both the polyclonal serum and mAb were used in immunofluorescence experiments, which showed binding of antibodies to the yeast cell surface. The mAb successfully opsonized P. lutzii and P. brasiliensis cells in co-incubations with J774.16 macrophage-like cells. Our results suggest that this mAb could serve as the basis for new immunotherapy regimens for PCM.

scholarly journals Monoclonal antibodies to different epitopes on a cell-surface enzyme, human placental alkaline phosphatase, effect different patterns of labeling with protein A-colloidal gold.

1987 ◽  
Vol 35 (11) ◽  
pp. 1277-1284 ◽  
Author(s):  
R Jemmerson ◽  
M Agre
Keyword(s):  
Alkaline Phosphatase ◽  
Monoclonal Antibodies ◽  
Cell Surface ◽  
Colloidal Gold ◽  
Polyclonal Antibodies ◽  
Protein A ◽  
Placental Alkaline Phosphatase ◽  
Gold Particles ◽  
Human Placental Alkaline Phosphatase ◽  
Membrane Components

Two monoclonal antibodies (mAbs) to different epitopes on human placental alkaline phosphatase (PLAP), both of the immunoglobulin G2a heavy-chain class and having similar affinities for PLAP, were compared for their ability to label the enzyme on the HeLa cell surface. In one type of experiment employing [125I]-labeled mAbs, the results demonstrated quantitative differences in binding of the mAbs to the cells. At saturating levels, the number of molecules of mAb E5 bound to the cells was almost eight times the number of mAb B10 molecules bound. In another type of experiment, mAbs were indirectly visualized on the cell surface using protein A tagged with colloidal gold particles in transmission electron microscopy. Only one of the antibodies (E5) displayed a clustered distribution of PLAP that previously had been observed with rabbit polyclonal antibodies and goat anti-rabbit IgG-labeled gold (J Histochem Cytochem 33:1227, 1985). The other antibody (B10) showed less frequent and more scattered labeling; three to four times more gold particles were visualized in each cluster on cells bound by mAb E5 compared to cells bound by B10. These results are consistent with the idea that not all epitopes on a membrane-bound antigen may be equally accessible for antibody binding. Even identical epitopes on different PLAP molecules are not equally hindered by other membrane components, since at least some of the PLAP molecules are labeled by the more sterically hindered mAb B10. Quantification of the number of gold particles employing the more abundantly bound mAb E5 provides an average estimate of seven to eight molecules of PLAP in each cluster. Because of inefficiencies in labeling, however, this value is probably lower than the real number.

scholarly journals Monoclonal Antibodies against SARS-CoV-2: Current Scenario and Future Perspectives

2021 ◽  
Vol 14 (12) ◽  
pp. 1272
Author(s):  
Eugenia Quiros-Roldan ◽  
Silvia Amadasi ◽  
Isabella Zanella ◽  
Melania Degli Antoni ◽  
Samuele Storti ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Therapeutic Potential ◽  
Severe Disease ◽  
Medical Field ◽  
Moderate Disease ◽  
Critical Issues ◽  
Risk Of Progression ◽  
Current Scenario ◽  
Manufacturing Techniques ◽  
Insight Into

Monoclonal antibodies (mAbs) have been known since the 1970s. However, their therapeutic potential in the medical field has recently emerged, with the advancement of manufacturing techniques. Initially exploited mainly in the oncology field, mAbs have become increasingly relevant in Infectious Diseases. Numerous mAbs have been developed against SARS-CoV 2 and have proven their effectiveness, especially in the management of the mild-to-moderate disease. In this review, we describe the monoclonal antibodies currently authorized for the treatment of the coronavirus disease 19 (COVID-19) and offer an insight into the clinical trials that led to their approval. We discuss the mechanisms of action and methods of administration as well as the prophylactic and therapeutic labelled indications (both in outpatient and hospital settings). Furthermore, we address the critical issues regarding mAbs, focusing on their effectiveness against the variants of concern (VoC) and their role now that a large part of the population has been vaccinated. The purpose is to offer the clinician an up-to-date overview of a therapeutic tool that could prove decisive in treating patients at high risk of progression to severe disease.

Expression Cassette and Plasmid Construction for Yeast Surface Display in Saccharomyces Cerevisiae

2021 ◽  
Author(s):  
Renan E A Piraine ◽  
Vitória S Gonçalves ◽  
Alceu GS dos Santos Junior ◽  
Rodrigo C Cunha ◽  
Pedro MM Albuquerque ◽  
...  
Keyword(s):  
Cell Surface ◽  
Recombinant Protein Expression ◽  
Polyclonal Antibodies ◽  
Surface Display ◽  
Cell Surface Display ◽  
Yeast Surface Display ◽  
Expression Cassette ◽  
Dot Blot ◽  
Herpesvirus Type ◽  
Yeast Cell Surface

Abstract Objectives. Develop a Cell Surface Display system in S. cerevisiae, based on the construction of an expression cassette for pYES2 plasmid. Results. The construction of an expression cassette containing the α-factor signal peptide and the C-terminal portion of the α-agglutinin protein was made and its sequence inserted into a plasmid named pYES2/gDαAgglutinin, allowing cell surface display of bovine herpesvirus type 5 (BoHV-5) glycoprotein D (gD) on S. cerevisiae BY4741 strain. Recombinant protein expression was confirmed by dot blot, and indirect immunofluorescence using monoclonal anti-histidine antibodies and polyclonal antibodies from mice experimentally vaccinated with a recombinant gD. Conclusions. These results demonstrate that the approach and plasmid used represent not only an effective system for immobilizing proteins on the yeast cell surface, as well as a platform for immunobiologicals development.

An Immunoradiometric Assay for Factor VIII Related Antigen (VIIIRAg) Using Two Monoclonal Antibodies-Comparison with Polyclonal Rabbit Antibodies for Use in von Willebrand’s Disease Diagnosis

1984 ◽  
Vol 52 (03) ◽  
pp. 250-252 ◽  
Author(s):  
Y Sultan ◽  
Ph Avner ◽  
P Maisonneuve ◽  
D Arnaud ◽  
Ch Jeanneau
Keyword(s):  
Monoclonal Antibodies ◽  
Structural Changes ◽  
Polyclonal Antibodies ◽  
Disease Diagnosis ◽  
Immunoradiometric Assay ◽  
Von Willebrand's Disease ◽  
Von Willebrand’S Disease ◽  
Structural Abnormalities ◽  
Antigenic Material ◽  
Von Willebrand

SummaryTwo monoclonal antibodies raised against FVIII/von Willebrand protein were used in an immunoradiometric assay (IRMA) to measure this antigen in normal plasma and plasma of patients with different forms of von Willebrand’s disease. The first antibody, an IgG1 was used to coat polystyrene tubes, the second one, an IgG2a, iodinated and used in the second step. Both antibodies inhibit ristocetin induced platelet agglutination and react strongly with platelets, megacaryocytes and endothelial cells. The IRMA test using these antibodies showed greater sensitivity than that using rabbit polyclonal anti VIIIRAg antibodies. A good correlation between the two tests was nevertheless found when VIIIRAg was measured in the majority of patient’s plasma. However 5 patients from 3 different families showed more antigenic material in the rabbit antibody IRMA than in the monoclonal antibody IRMA. It is suggested therefore that the monoclonal antibodies identify part of the VIIIR:Ag molecule showing structural abnormalities in these vWd patients, these structural changes remaining undetected by the polyclonal antibodies.

Use of monoclonal and polyclonal antibodies to detect prostatic acid phosphatase by immunoblotting.

1986 ◽  
Vol 32 (10) ◽  
pp. 1832-1835 ◽  
Author(s):  
P C Patel ◽  
L Aubin ◽  
J Côte
Keyword(s):  
Monoclonal Antibodies ◽  
Acid Phosphatase ◽  
Western Blot ◽  
Polyclonal Antibodies ◽  
Prostatic Acid Phosphatase ◽  
The Other ◽  
Dot Blot ◽  
Western Blots ◽  
Dot Blot Assay ◽  
Polyclonal Antisera

Abstract We investigated two techniques of immunoblotting--the Western blot and the dot blot--for use in detecting prostatic acid phosphatase (PAP, EC 3.1.3.2). We used polyclonal antisera to human PAP, produced in rabbits by hyperimmunization with purified PAP, and PAP-specific monoclonal antibodies in the immunoenzymatic protocols. We conclude that PAP can be readily detected by Western blots with use of polyclonal antisera, but not with monoclonal antibodies. On the other hand, using a dot blot assay, we could easily detect PAP with both polyclonal and monoclonal antibodies.

scholarly journals Effective use of monoclonal antibodies for treatment of persistent COVID-19 infection in a patient on rituximab

2021 ◽  
Vol 14 (8) ◽  
pp. e243469
Author(s):  
Carlos X Rabascall ◽  
Becky X Lou ◽  
Brianne Navetta-Modrov ◽  
Stella S Hahn
Keyword(s):  
Monoclonal Antibodies ◽  
Symptom Onset ◽  
Severe Disease ◽  
Therapeutic Window ◽  
Immunosuppressed Patient ◽  
Unique Case ◽  
Risk Of Progression ◽  
Immunosuppressed Patients ◽  
Effective Use ◽  
Potential Use

As we are over a year into the COVID-19 pandemic, we have made many forward strides in therapeutics. These treatments, such as monoclonal antibodies, have help mitigate the detrimental and often fatal consequences of COVID-19. The current indication for the use of monoclonal antibodies is mild to moderate COVID-19 infection within 10 days of symptom onset in those who are at high risk of progression to severe disease. However, their role in patients with prolonged symptoms is not clear. We present a unique case of monoclonal antibodies use after 54 days of symptom onset in an immunosuppressed patient with persistent COVID-19 infection despite standard treatment. This case illustrates the potential use of monoclonal antibodies outside of the current recommended therapeutic window in immunosuppressed patients, who may have difficulty with viral clearance.

scholarly journals Paracoccidioides brasiliensis Isolated from Nine-Banded Armadillos (Dasypus novemcinctus) Reveal Population Structure and Admixture in the Amazon Basin

2021 ◽  
Vol 7 (1) ◽  
pp. 54
Author(s):  
Eduardo Bagagli ◽  
Daniel Ricardo Matute ◽  
Hans Garcia Garces ◽  
Bernardo Guerra Tenório ◽  
Adalberto Garcia Garces ◽  
...  
Keyword(s):  
Population Structure ◽  
Latin America ◽  
Endemic Area ◽  
Amazon Basin ◽  
Paracoccidioides Brasiliensis ◽  
Pathogenic Fungi ◽  
Sensu Stricto ◽  
Species Level ◽  
Dasypus Novemcinctus ◽  
Show Evidence

Paracoccidioidomycosis is an endemic fungal disease to Latin America caused by at least five species-level genotypes of Paracoccidioides, named P. lutzii, P. brasiliensis (S1a and S1b populations), P. americana, P. restrepiensis, and P. venezuelensis. In this manuscript, we report on Paracoccidioides sp. sampling efforts in armadillos from two different areas in Brazil. We sequenced the genomes of seven Paracoccidioides isolates and used phylogenomics and populations genetics for genotyping. We found that P. brasiliensis and P. lutzii are both present in the Amazon region. Additionally, we identified two Paracoccidioides isolates that seem to be the result of admixture between divergent populations within P. brasiliensis sensu stricto. Both of these isolates were recovered from armadillos in a P. lutzii endemic area in Midwestern Brazil. Additionally, two isolates from human patients also show evidence of resulting from admixture. Our results suggest that the populations of P. brasiliensis sensu stricto exchange genes in nature. More generally, they suggest that population structure and admixture within species is an important source of variation for pathogenic fungi.

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