Identification of B-cell linear epitopes in domains 1–3 of pyolysin of Trueperella pyogenes using polyclonal antibodies

ABSTRACT The neutralizing activities of polyclonal antibodies and monoclonal antibodies (MAbs) obtained by immunization of mice with L1 virus-like particles (VLPs) were investigated by using pseudovirion infectivity assays for human papillomavirus type 16 (HPV-16), HPV-31, HPV-33, HPV-45, HPV-58, and HPV-59 to obtain a better definition of cross-neutralization between high-risk HPVs. In this study, we confirmed and extended previous studies indicating that most genital HPV genotypes represent separate serotypes, and the results suggest that the classification of serotypes is similar to that of genotypes. In addition, three cross-neutralizing MAbs were identified (HPV-16.J4, HPV-16.I23, and HPV-33.E12). MAb HPV-16.J4 recognized a conserved linear epitope located within the FG loop of the L1 protein, and HPV-16.I23 recognized another located within the DE loop. The results suggested that reactivity of MAb HPV-16.I23 to L1 protein is lost when leucine 152 of the HPV-16 L1 protein is replaced by phenylalanine. This confirmed the existence of linear epitopes within the L1 protein that induce neutralizing antibodies, and this is the first evidence that such linear epitopes induce cross-neutralization. However, the cross-neutralization induced by L1 VLPs represents less than 1% of the neutralizing activity induced by the dominant conformational epitopes, and it is questionable whether this is sufficient to offer cross-protection in vivo.

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Antibody Prophylaxis and Therapy against West NileVirus Infection in Wild-Type and ImmunodeficientMice

Journal of Virology ◽

10.1128/jvi.77.24.12941-12949.2003 ◽

2003 ◽

Vol 77 (24) ◽

pp. 12941-12949 ◽

Cited By ~ 134

Author(s):

Michael J. Engle ◽

Michael S. Diamond

Keyword(s):

B Cell ◽

Gamma Globulin ◽

Polyclonal Antibodies ◽

Current Treatment ◽

Mouse Serum ◽

Wild Type ◽

Viral Reservoirs ◽

In The Wild ◽

Human Gamma Globulin ◽

Immune Antibody

ABSTRACT West Nile virus (WNV) is a mosquito-borne Flavivirus that causes encephalitis in a subset of susceptible humans. Current treatment for WNV infections is supportive, and no specific therapy or vaccine is available. In this study, we directly tested the prophylactic and therapeutic efficacy of polyclonal antibodies against WNV. Passive administration of human gamma globulin or mouse serum prior to WNV infection protected congenic wild-type, B-cell-deficient (μMT), and T- and B-cell-deficient (RAG1) C57BL/6J mice. Notably, no increased mortality due to immune enhancement was observed. Although immune antibody completely prevented morbidity and mortality in wild-type mice, its effect was not durable in immunocompromised mice: many μMT and RAG1 mice eventually succumbed to infection. Thus, antibody by itself did not completely eliminate viral reservoirs in host tissues, consistent with an intact cellular immune response being required for viral clearance. In therapeutic postexposure studies, human gamma globulin partially protected against WNV-induced mortality. In μMT mice, therapy had to be initiated within 2 days of infection to gain a survival benefit, whereas in the wild-type mice, therapy even 5 days after infection reduced mortality. This time point is significant because between days 4 and 5, WNV was detected in the brains of infected mice. Thus, passive transfer of immune antibody improves clinical outcome even after WNV has disseminated into the central nervous system.

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Mapping and characterization of B cell linear epitopes in the conservative regions of hepatitis C virus envelope glycoproteins

Journal of Viral Hepatitis ◽

10.1046/j.1365-2893.2002.00358.x ◽

2002 ◽

Vol 9 (3) ◽

pp. 174-182 ◽

Cited By ~ 13

Author(s):

L. V. Olenina ◽

L. I. Nikolaeva ◽

B. N. Sobolev ◽

N. P. Blokhina ◽

A. I. Archakov ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

B Cell ◽

Envelope Glycoproteins ◽

Virus Envelope ◽

Linear Epitopes

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Ontogeny of immunoglobulin expression in the Mexican axolotl

Development ◽

10.1242/dev.107.2.253 ◽

1989 ◽

Vol 107 (2) ◽

pp. 253-263

Author(s):

J.S. Fellah ◽

D. Vaulot ◽

A. Tournefier ◽

J. Charlemagne

Keyword(s):

B Cell ◽

Polyclonal Antibodies ◽

Relative Proportion ◽

Absolute Number ◽

Ambystoma Mexicanum ◽

Temporal Expression ◽

Cell Compartment ◽

Mexican Axolotl ◽

Marked Enhancement ◽

Ig Synthesis

The ontogeny of immunoglobulin (Ig) synthesis was followed at both cellular and serological levels in the Mexican axolotl (Ambystoma mexicanum) using polyclonal antibodies recognizing all Ig molecules and a set of monoclonal antibodies (Mabs) specific for the C mu and Cv heavy Ig chain isotypes and for the light chain constituents shared by IgM and IgY molecules. Clusters of IgM- and of IgY-synthesizing lymphocytes, often located in separate sites, are first present in spleen sections of 7-week-old 25 mm larvae, about one month after differentiation of the spleen anlage (stage 39–40). In 12-week-old 30–35 mm larvae, the relative proportion of IgM- and IgY-synthesizing cells in the spleen is the same as that in adult animals. However, a marked enhancement of the spleen B cell compartment occurs from 5 to 9 months when Ig-positive cells represent about 88% of the lymphocytes population compared to 60% in adults. No structures equivalent to B cell germinal centers were observed at any stage of the spleen differentiation and cells, although often clustered in small groups, remain dispersed in the entire organ. The relative proportions of IgM and IgY B cells throughout the spleen remain constant during development (about 1 IgY+ cell for 5–6 IgM+ cells) and IgM molecules are first detected in the serum of 2.5-month-old larvae. The enhancement of the serum IgM level correlates well with the absolute number of IgM+ cells in the growing spleen. IgY molecules cannot be detected in the serum before the 7th month but their level quickly increases to reach about 60% of the adult value at 10 months. Thyroxine-induced metamorphosis or hyperimmunization of 4- to 6-month-old larvae had no effect upon the temporal expression of the Ig classes in serum.

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B-Cell Linear Epitopes in the Conservative Regions of Hepatitis C Virus Envelope Glycoproteins

Peptides: The Wave of the Future ◽

10.1007/978-94-010-0464-0_482 ◽

2001 ◽

pp. 1031-1032

Author(s):

Ekaterina F. Kolesanova ◽

Ludmila V. Olenina ◽

Boris N. Sobolev ◽

Ludmila I. Nikolaeva ◽

Alexander I. Archakov

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

B Cell ◽

Envelope Glycoproteins ◽

Virus Envelope ◽

Linear Epitopes

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Rational design of peptides for identification of linear epitopes and generation of neutralizing monoclonal antibodies against DKK2 for cancer therapy

Antibody Therapeutics ◽

10.1093/abt/tbaa004 ◽

2020 ◽

Vol 3 (2) ◽

pp. 63-70 ◽

Cited By ~ 1

Author(s):

Rongqing Zhao ◽

Qian Xiao ◽

Maohua Li ◽

Wenlin Ren ◽

Chenxi Xia ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Tumor Growth ◽

Wnt Signaling ◽

Rational Design ◽

Polyclonal Antibodies ◽

Wnt Signaling Pathway ◽

Keyhole Limpet Hemocyanin ◽

Linear Epitopes

Abstract Dickkopf-related protein 2 (DKK2)is a member of the Dickkopf family in Wnt signaling pathway. Recently, we found that antibodies against DKK2 could activate natural killer (NK) and CD8+ T cells in tumors and inhibit tumor growth. In this paper, we report the rational design of peptides for identification of linear epitopes and generation of neutralizing monoclonal anti-DKK2 antibodies. To break the immune tolerance, we designed and chemically synthesized six peptides corresponding to different regions of DKK2 as immunogens and found five of them could generate mouse polyclonal antibodies that can bind to the active recombinant human DKK2 protein. Neutralizing mouse monoclonal antibodies (5F8 and 1A10) against human DKK2 were successfully developed by immunizing the mice with two different peptides (34KLNSIKSSL42 and 240KVWKDATYS248) conjugated to Keyhole limpet hemocyanin (KLH). The monoclonal antibodies not only abolish DKK2’s suppression of Wnt signaling in vitro but also inhibits tumor growth in vivo. Currently, those two mAbs are undergoing humanization as immunotherapy candidates and may offer a new drug for treatment of human cancers.

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Identification of specific B cell linear epitopes of mycoplasma hyorhinis P37 protein using monoclonal antibodies against baculovirus-expressed P37 protein

BMC Microbiology ◽

10.1186/s12866-019-1614-4 ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 1

Author(s):

Hongzhen Zhu ◽

Yanwu Wei ◽

Liping Huang ◽

Dan Liu ◽

Yongxing Xie ◽

...

Keyword(s):

Amino Acids ◽

Monoclonal Antibodies ◽

B Cell ◽

Etiologic Agent ◽

Mycoplasma Hyorhinis ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Precise Analysis ◽

Linear Epitopes ◽

Specific Antigens

Abstract Background Mycoplasma hyorhinis (Mhr) is the etiologic agent of lameness and polyserositis in swine. P37 is a membrane protein of Mhr that may be an important immunogen and is a potential target for diagnostic development. However, there is little information concerning Mhr P37 protein epitopes. A precise analysis of the P37 protein epitopes should extend our understanding of the antigenic composition of the P37 protein and the humoral immune responses to Mhr infection. Investigating the epitopes of Mhr P37 will help to establish a detection method for Mhr in tissue and provide an effective tool for detecting Mhr infection. Results Western blot and indirect immunofluorescence assays (IFA) confirmed that the expressed P37 protein was recognized by Mhr-positive porcine and mouse sera. Furthermore, the P37 protein was purified using affinity chromatography and used to immunize mice for hybridoma cell fusion. Four monoclonal antibodies (mAbs) found to be positive for Mhr were detected in infected lung tissue. A panel of truncated P37 proteins was used to identify the minimal B cell linear epitopes of the protein based on these mAbs. The core epitope was determined to be 206KIKKAWNDKDWNTFRNF222. Conclusions In this study, we identified 17 critical amino acids that determine the epitope of the P37 protein of Mhr. This study identified mAbs that could provide useful tools for investigating the Mhr P37 antigenic core epitope (amino acids 206–222) and detecting Mhr-specific antigens in infected tissue.

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Monoclonal Antibodies Targeting the HR2 Domain and the Region Immediately Upstream of the HR2 of the S Protein Neutralize In Vitro Infection of Severe Acute Respiratory Syndrome Coronavirus

Journal of Virology ◽

10.1128/jvi.80.2.941-950.2006 ◽

2006 ◽

Vol 80 (2) ◽

pp. 941-950 ◽

Cited By ~ 41

Author(s):

Kuo-Ming Lip ◽

Shuo Shen ◽

Xiaoming Yang ◽

Choong-Tat Keng ◽

Aihua Zhang ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Polyclonal Antibodies ◽

Heptad Repeat ◽

Functional Domain ◽

Protein Fragment ◽

S Protein ◽

Linear Epitopes ◽

Respiratory Coronavirus ◽

Neutralizing Epitopes

ABSTRACT We have previously shown that an Escherichia coli-expressed, denatured spike (S) protein fragment of the severe acute respiratory coronavirus, containing residues 1029 to 1192 which include the heptad repeat 2 (HR2) domain, was able to induce neutralizing polyclonal antibodies (C. T. Keng, A. Zhang, S. Shen, K. M. Lip, B. C. Fielding, T. H. Tan, C. F. Chou, C. B. Loh, S. Wang, J. Fu, X. Yang, S. G. Lim, W. Hong, and Y. J. Tan, J. Virol. 79:3289-3296, 2005). In this study, monoclonal antibodies (MAbs) were raised against this fragment to identify the linear neutralizing epitopes in the functional domain and to investigate the mechanisms involved in neutralization. Eighteen hybridomas secreting the S protein-specific MAbs were obtained. Binding sites of these MAbs were mapped to four linear epitopes. Two of them were located within the HR2 region and two immediately upstream of the HR2 domain. MAbs targeting these epitopes showed in vitro neutralizing activities and were able to inhibit cell-cell membrane fusion. These results provide evidence of novel neutralizing epitopes that are located in the HR2 domain and the spacer region immediately upstream of the HR2 of the S protein.

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Histologic Classification and Immunophenotype of Lymphosarcomas in Cats with Naturally and Experimentally Acquired Feline Immunodeficiency Virus Infections

Veterinary Pathology ◽

10.1177/030098589603300302 ◽

1996 ◽

Vol 33 (3) ◽

pp. 264-272 ◽

Cited By ~ 75

Author(s):

J. J. Callanan ◽

B. A. Jones ◽

J. Irvine ◽

B. J. Willett ◽

I. A. P. McCandlish ◽

...

Keyword(s):

B Cell ◽

Feline Immunodeficiency Virus ◽

Cell Activation ◽

Polyclonal Antibodies ◽

Leukemia Virus ◽

Feline Leukemia Virus ◽

B Cell Activation ◽

Virus Infections ◽

Histologic Classification ◽

Immunodeficiency Virus

Lymphosarcoma (malignant lymphoma) is the commonest hematopoietic tumor in the cat. Many cases are associated with feline leukemia virus (FeLV) infection, but epidemiologic and experimental data suggest that feline immunodeficiency virus (FIV) may also have a role in lymphomagenesis. In this paper, we describe the clinical presentation, histologic classification, and immunophenotype of lymphosarcoma in eight domestic cats with natural or experimental FIV infections. The tumors were often of unusual distribution, with the majority of cases conforming to the least common anatomic classification of “miscellaneous.” Histopathologic and immunophenotypic analysis using a panel of anti-cat and cross-reactive anti-human monoclonal and polyclonal antibodies identified seven of these tumors as high-grade B cell lymphomas of the centroblastic or immunoblastic subtypes. The remaining case was a T-cell tumor associated with a concurrent FeLV infection. Our findings, together with the results of an analysis of FIV proviral DNA in these tumors, indicate that the B-cell lymphosarcomas were comparable to those observed in human and simian immunodeficiency virus infections and that the role of FIV in lymphomagenesis is indirect and related to the potential for malignant transformation during polyclonal B cell activation.

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Antibody Mapping of the Linear Epitopes of CMY-2 and SHV-1 β-Lactamases

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.10.3980-3988.2004 ◽

2004 ◽

Vol 48 (10) ◽

pp. 3980-3988 ◽

Cited By ~ 15

Author(s):

Andrea M. Hujer ◽

Christopher R. Bethel ◽

Robert A. Bonomo

Keyword(s):

Amino Acids ◽

Polyclonal Antibodies ◽

Synthesis Method ◽

Hydrolytic Activity ◽

Spot Synthesis ◽

Antibody Recognition ◽

Molar Excess ◽

Omega Loop ◽

Direct Binding ◽

Linear Epitopes

ABSTRACT Knowledge of the amino acids that define recognition of anti-β-lactamase antibodies is critical to the interpretation of sensitivity and specificity of these antibodies when they are used in a clinical or research setting. To this end, we mapped the epitopes of the CMY-2 and SHV-1 β-lactamases by using the SPOT synthesis method. Eight linear epitopes in SHV-1 and seven linear epitopes in CMY-2 were identified by using anti-SHV-1 and anti-CMY-2 polyclonal antibodies, respectively. The epitopes of SHV-1 were mapped to amino acids at the Ambler positions ABL 28 to 38, 42 to 54, 88 to 100, 102 to 114, 170 to 182, 186 to 194, 202 to 210, and 276 to 288. In the epitope spanning amino acids 102 to 114, alanine and X-Scan analysis demonstrated that D104, Y105, P107, and S109 are essential residues for antibody recognition. In the epitope containing amino acids 170 to 182, N170, L173, P174, G175, and D176 were immunodominant. In CMY-2 β-lactamase, amino acids 4 to 16, 70 to 79, 211 to 223, 274 to 286, 289 to 298, 322 to 334, and 343 to 358 of the mature enzyme defined the major linear epitopes. A detailed analysis of the recognition sites that are located in an area analogous to the omega loop of class A β-lactamases (V211 to V223) showed that the amino acids Q215 to E219 are important in antibody binding. Incubation of CMY-2 β-lactamase with a 10-fold molar excess of anti-CMY-2 antibody for 60 min resulted in greater than 80% inhibition of nitrocefin hydrolysis. A 10-fold molar excess of anti-SHV-1 antibody reduced the activity of SHV-1 by 69%. Analysis of the CMY-2 and SHV-1 structures suggest that this reduction of hydrolytic activity may be due in part to the direct binding of antibodies to the omega loop, thereby hindering access of substrate to the active site.

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