scholarly journals Circumsporozoite proteins of malaria parasites contain a single immunodominant region with two or more identical epitopes.

1983 ◽  
Vol 157 (6) ◽  
pp. 1947-1957 ◽  
Author(s):  
F Zavala ◽  
A H Cochrane ◽  
E H Nardin ◽  
R S Nussenzweig ◽  
V Nussenzweig
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Specific Binding ◽  
Polyclonal Antibodies ◽  
Binding Assays ◽  
Malaria Parasites ◽  
Immunodominant Region ◽  
Additional Finding ◽  
Single Region ◽  
Competition Binding

We have used panels of monoclonal antibodies to circumsporozoite (CS) proteins of Plasmodium falciparium, P. vivax, and P. knowlesi to determine the number of topographically independent epitopes of these antigens. The results of competition binding assays indicated that single regions of the CS molecules were recognized by the homologous monoclonal antibodies. Competition binding assays were also used to study the specificity of antibodies contained in the sera of humans and monkeys that had developed sterile immunity after immunization with irradiated, intact sporozoites. We found that single monoclonal antibodies inhibited 70-95% of the specific binding of the polyclonal antibodies to crude extracts of sporozoites. It appears, therefore, that CS proteins are among the most immunogenic constituents of sporozoites, and that a single region of these molecules contains most of the immunogenic activity. An additional finding was that the immunodominant region of CS molecules is multivalent with regard to the expression of a single epitope. This was demonstrated by the ability of monomers of CS proteins to bind simultaneously two or more molecules of the same monoclonal antibody.

Reactions of human antibodies with CR1 immobilised by mouse monoclonal antibody E11 Red cell phenotype Murine MAb Human anti- Absorbance Ratio Kn(a+) ] Kn(a-) E11 Kna 0.755 0.195 4:1 McC(a+) 0.538 McC(a-) E11 McC 0.136 4:1 Yk(a+) 0.315 Yk(a-) E11 Yka 0.120 26:1 Sl(a+) 0.342 Sl(a-) E11 Sla 0.074 4.6:1 Cs(a+) 0.139 Cs(a-) E11 Cs 0.108 Mapping relative positions of antigens on a specific protein When several murine monoclonal antibodies to different epitopes on the same protein are available, MAIEA can be used to study the relative position of antigens on that protein. This application of MAIEA depends on mutual inhibition of murine monoclonal antibodies and human antibodies. A negative result is obtained when human and monoclonal antibodies compete for the same epitope, or bind to very closely located epitopes, so no tri-molecular complex is produced. Several monoclonal antibodies to the Kell protein have been used in MAIEA to study the relationships of the Kell system antigens [10]. The decay accelerating factor DAF, CD55, is detected by several monoclonal antibodies. Three antibodies BRIC 230, BRIC 110 and BRIC 216 were known from competitive binding assays to bind to different short consensus repeats (SCR) [11]. So three of the four SCRs of the DAF molecule were positively identified (Table II). Strong positive reactions were observed with all three BRIC antibodies and anti-Cr3, anti-WES8, and anti-WESb showing that MAIEA is a useful techique for studying this system [12]. The results showed that Cr8, WESa, and WESb are not on the first three SCRs and must

1995 ◽  
pp. 190-190
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Molecular Complex ◽  
Specific Protein ◽  
Cell Phenotype ◽  
Binding Assays ◽  
Human Antibodies ◽  
Short Consensus Repeats ◽  
Murine Monoclonal Antibodies ◽  
Competitive Binding Assays

scholarly journals Functional study of a monoclonal antibody to IgE Fc receptor (Fc epsilon R2) of eosinophils, platelets, and macrophages.

1986 ◽  
Vol 164 (1) ◽  
pp. 72-89 ◽  
Author(s):  
M Capron ◽  
T Jouault ◽  
L Prin ◽  
M Joseph ◽  
J C Ameisen ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
B Lymphocyte ◽  
Specific Binding ◽  
Polyclonal Antibodies ◽  
Fc Receptor ◽  
The Other ◽  
Functional Study ◽  
Low Density ◽  
Fluorescence Staining ◽  
Ige Binding

An IgM mAb (BB10) was produced by immunization of mice with human eosinophils purified according to their abnormal low density ("hypodense" cells), and previously shown to exhibit increased IgE-dependent antiparasite cytotoxicity. This BB10 antibody, selected for positive fluorescence staining of hypodense blood or lung eosinophils and low or negative staining of normodense eosinophils or neutrophils, could strongly inhibit IgE-dependent cytotoxicity of human eosinophils and platelets. The specificity for the IgE Fc receptor was suggested by the high levels of inhibition of IgE rosettes formed by eosinophils after incubation with the purified IgM fraction of BB10, whereas other receptors (Fc gamma R, CR1) were not affected. On the other hand, BB10, able to inhibit rat eosinophil Fc epsilon R, did not react with the IgE Fc receptor on mast cells or basophils. A technique using radioiodinated BB10 allowed us to quantify the specific binding of BB10 to human eosinophils and platelets. Competition experiments revealed a crossinhibition between the binding of BB10 and IgE, suggesting the specificity of BB10 for the IgE binding site of eosinophil, platelet, and monocyte Fc epsilon R. Three proteins having extrapolated Mr of 32,000, 43,000-45,000, and 97,000 were found in the platelet extract eluted from a BB10 or from an IgE immunosorbent column. These findings confirm the similarities between IgE Fc receptors on human eosinophils, platelets, and macrophages, already observed with polyclonal antibodies directed against the B lymphocyte Fc epsilon receptor. They suggest, moreover, that the mAb BB10 can represent a good reagent for further investigations on the structure and the functions of this IgE Fc receptor (Fc epsilon R2).

Abstract 14697: Novel Assays for Quantification of Lipoprotein-Associated (PCSK9-apoB, PCSK9-Lp(a)) Proprotein Covertase Subtilisin/Kexin Type 9 (PCKS9)

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Calvin Yeang ◽  
Yun-Seok Choi ◽  
Sang-Rok Lee ◽  
Monica L Bertoia ◽  
Eric B Rimm ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Human Plasma ◽  
Polyclonal Antibodies ◽  
Human Monoclonal Antibody ◽  
Ldl Receptor ◽  
Plasma Lipoproteins ◽  
Health Study ◽  
Clinical Samples ◽  
Ldl Particles

Background: PCSK9 is a major regulator of plasma LDL-C. Monoclonal antibodies to PCSK9 lower LDL-C by 45-65% and Lp(a) by 9-38%. The canonical function of PCSK9 is binding of LDL-receptor (LDLR) via its extracellular EGF-A domain, and subsequently mediating LDLR degradation. However, PCSK9 also weakly associates with plasma lipoproteins, with 20-40% of total plasma PCSK9 found on LDL. However, most LDL particles do not contain PCSK9. Whether PCSK9 also associates with other lipoproteins such as Lp(a) are not well described. Methods: Sensitive and quantitative sandwich-based ELISA assays were developed to measure PCSK9 associated plasma lipoproteins in both mouse and human plasma. For human plasma, commercial rabbit polyclonal antibodies binding to the C-terminal region of PCSK9 (Abgent, ThermoFisher) or REGN727 human monoclonal antibody were bound to microtiter well plates. Plasma was added and monoclonal antibodies MB47 and LPA4, binding to apoB-100 and apo(a) respectively, were used to detect PCSK9-apoB-100 and PCSK9-Lp(a) complexes with a chemiluminescent ELISA. For mouse assays, REGN727 was used as the capture antibody as it detects mouse PCSK9 and monoclonal antibody LF3 was used to detect mouse apoB. Results: PCSK9-apoB and PCSK9-Lp(a) complexes could be detected in both human plasma and in various mouse models expressing apo(a) or Lp(a). The signal to noise ratio was ~20 fold in various clinical samples, including in healthy subjects and in patients with cardiovascular disease. In 536 clinical samples from the Health Professional Follow-Up Study, PCSK9-Lp(a) correlated strongly with Lp(a) (r=0.59, p<0.001, age-adjusted) but not other lipid variables. PCSK9-apoB correlated weakly with PCSK9-Lp(a) (r=0.30, p<0.001, age-adjusted) and LDL-C (r=0.22, p<0.001, age-adjusted). These associations were virtually the same in 526 women in the Nurses’ Health Study. Conclusions: Novel ELISAs were generated to quantitate lipoprotein-associated PCSK9 in transgenic mouse and human plasma, including on apoB and Lp(a). Changes in PCSK9-Lp(a) complexes may provide insights into the Lp(a)-lowering effect of PCSK9 antibodies. Whether these assays will predict CVD outcomes waits to be determined in PCSK9 antibody and epidemiological studies.

scholarly journals A comparative study of the use of monoclonal antibodies using three different immunohistochemical methods: an evaluation of monoclonal and polyclonal antibodies against human prostatic acid phosphatase.

1982 ◽  
Vol 30 (3) ◽  
pp. 253-260 ◽  
Author(s):  
W Y Naritoku ◽  
C R Taylor
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Acid Phosphatase ◽  
Polyclonal Antibodies ◽  
Prostatic Acid Phosphatase ◽  
Mouse Serum ◽  
Paraffin Sections ◽  
Layer System ◽  
Significant Difference ◽  
Immunohistochemical Methods

The use of immunohistochemical methods has been advocated for the detection and localization of prostatic acid phosphatase in paraffin sections of human prostate. This article explores the possible advantages of utilizing monoclonal antibodies in this method. Monoclonal antibodies, specific for human prostatic acid phosphatase, were integrated into three different immunohistochemical procedures. In the first method, a three-layer peroxidase-antiperoxidase (PAP) system was employed; the monoclonal antibody was followed by rabbit bridge antibody directed against mouse immunoglobulin and mouse PAP complex. The second method was a three-layer system utilizing biotin-labeled horse anti-mouse antibody as "bridge" antiserum between the primary monoclonal antibody and an avidin-biotin-horseradish peroxidase complex. The third method was a four-layer system; the monoclonal antibody was followed by rabbit anti-mouse serum, swine anti-rabbit immunoglobulin as the bridge antibody and rabbit PAP complex. It was found that some, but not all, monoclonal antibodies can be used for the detection of prostatic acid phosphatase in paraffin sections. The four-layer PAP method was found to be the most sensitive method of the three systems tested; however, the avidin-biotin method required the least amount of time. No significant difference in the quality of staining was observed between monoclonal antibodies and carefully absorbed conventional antiserum.

scholarly journals Identification of Residues within Human Glycoprotein VI Involved in the Binding to Collagen

2004 ◽  
Vol 279 (50) ◽  
pp. 52293-52299 ◽  
Author(s):  
Christelle Lecut ◽  
Véronique Arocas ◽  
Hans Ulrichts ◽  
Anthony Elbaz ◽  
Jean-Luc Villeval ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Molecular Modeling ◽  
Binding Sites ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Binding Assays ◽  
Glycoprotein Vi ◽  
Ligand Binding Sites ◽  
Human Receptor

Glycoprotein VI (GPVI) has a crucial role in platelet responses to collagen. Still, little is known about its interaction with its ligands. In binding assays using soluble or cell-expressed human GPVI, we observed that (i) collagen, and the GPVI-specific ligands collagen-related peptides (CRP) and convulxin, competed with one another for the binding to GPVI and (ii) monoclonal antibodies directed against the extracellular part of the human receptor displayed selective inhibitory properties on GPVI interaction with its ligands. Monoclonal antibody 9E18 strongly reduced the binding of GPVI to collagen/CRP, 3F8 inhibited its interaction with convulxin, whereas 9O12 prevented all three interactions. These observations suggest that ligand-binding sites are distinct, exhibiting specific features but at the same time also sharing some common residues participating in the recognition of these ligands. The epitope of 9O12 was mapped by phage display, along with molecular modeling of human GPVI, which allowed the identification of residues within GPVI potentially involved in ligand recognition. Site-directed mutagenesis revealed that valine 34 and leucine 36 are critical for GPVI interaction with collagen and CRP. The loop might thus be part of a collagen/CRP-binding site.

Production of Monoclonal Antibodies Against Enamelin and Against Amelogenin Proteins of Developing Enamel Matrix

1987 ◽  
Vol 1 (2) ◽  
pp. 282-288 ◽  
Author(s):  
D. Deutsch ◽  
A. Palmon ◽  
J. Catalano-Sherman ◽  
R. Laskov
Keyword(s):  
Monoclonal Antibody ◽  
Extracellular Matrix ◽  
Monoclonal Antibodies ◽  
Structural Organization ◽  
Polyclonal Antibodies ◽  
Protein Band ◽  
Enamel Matrix ◽  
Complex Process ◽  
The One ◽  
Successful Production

The extracellular matrix of developing enamel contains two major classes of proteins, the hydrophobic proline-rich amelogenins and the acidic serine-, glycine-, and aspartic-rich enamelins. These proteins have been postulated as playing a major role in the mineralization and structural organization of developing enamel. To identify and further characterize these different proteins and their possible role in this complex process of biological mineralization, we have in recent years been concerned with the production of specific probes for these proteins. Previously, we have reported on the successful production of specific polyclonal antibodies against enamelin proteins, which did not cross-react with amelogenins, and against amelogenin proteins, which did not cross-react with enamelins (Deutsch et al., 1986, 1987). We now report the production of monoclonal antibodies against a major bovine amelogenin protein (28 kDa) and against a major bovine enamelin protein (66 kDa). One monoclonal antibody against amelogenin and one against enamelin are described. The results showed that the monoclonal antibody against the amelogenin protein reacted strongly with the 28-kDa amelogenin protein band but did not cross-react with enamelins, and the one against the enamelin protein reacted with the 66-kDa enamelin protein but did not cross-react with amelogenins. These monoclonal antibodies provide a specific and powerful tool to distinguish between and further characterize these different classes of proteins, and to improve our understanding of the process of enamel formation.

Competitive Direct Enzyme-Linked Immunosorbent Screening Assay for the Detection of Sulfamethazine Contamination of Animal Feeds

1991 ◽  
Vol 74 (5) ◽  
pp. 784-789 ◽  
Author(s):  
Dixon E Holland Deborah ◽  
Stanley E Katz
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Polyclonal Antibodies ◽  
Screening Method ◽  
Screening Assay ◽  
Animal Feeds ◽  
Feed Samples

Abstract A sensitive screening method has been developed for detecting sulfamethazine (SMZ) contamination of feeds by using either polyclonal or monoclonal antibodies and a direct competitive enzyme-linked immunosorbent screening assay (ELISA). Feed samples of 25.0 g are extracted with 0.5N HCI and centrifuged. The extract is adjusted to pH 7.0 with 3.0N NaOH and recentrifuged. This pH-adjusted extract is used in the EUSA. Levels as low as 0.004 μg SMZ/g feed were detected In supplemented extracts by polyclonal antibodies; levels of 0.4 μg SMZ/g feed were detected by a monoclonal antibody.

scholarly journals Circumsporozoite proteins of human malaria parasites Plasmodium falciparum and Plasmodium vivax.

1982 ◽  
Vol 156 (1) ◽  
pp. 20-30 ◽  
Author(s):  
E H Nardin ◽  
V Nussenzweig ◽  
R S Nussenzweig ◽  
W E Collins ◽  
K T Harinasuta ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Plasmodium Falciparum ◽  
Monoclonal Antibodies ◽  
Plasmodium Vivax ◽  
Indirect Immunofluorescence ◽  
Immune Protection ◽  
Entire Surface ◽  
Malaria Parasites ◽  
Precipitin Reaction ◽  
Human Volunteers

Monoclonal antibodies were raised against sporozoites of two species of malaria parasites, Plasmodium falciparum and Plasmodium vivax. The antibodies reacted with polypeptides (circumsporozoite proteins) that are uniformly distributed over the entire surface of sporozoites, as shown by indirect immunofluorescence and by the circumsporozoite precipitin reaction. The epitopes recognized by the monoclonal antibodies were expressed on sporozoites from different geographical isolates of the homologous species but were not detected on sporozoites of heterologous species nor on blood forms of the parasite. The monoclonal antibody to P. falciparum specifically immunoprecipitated two polypeptides of apparent 67,000 mol wt (Pf67) and 58,000 mol wt (Pf58) from extracts of [35S]methionine-labeled P. falciparum sporozoites. Similarly, the anti-P. vivax monoclonal immunoprecipitated two proteins of 51,000 mol wt (Pv51) and 45,000 mol wt (Pv45) from extracts of metabolically labeled P. vivax sporozoites. The extracts were also reacted with the serum of human volunteers successfully vaccinated with sporozoites of either P. vivax or P. falciparum. The patterns of immunoprecipitation were almost identical to those obtained with the corresponding monoclonal antibodies. The circumsporozoite proteins of P. falciparum and P. vivax play a role in immune protection. Incubation of the appropriate monoclonal antibody with viable sporozoites of the homologous species significantly reduced parasite infectivity, as determined by sporozoite neutralization assays carried out in splenectomized chimpanzees.

Interaction of monoclonal antibodies with growth hormone-binding protein and its complex with growth hormone

1993 ◽  
Vol 139 (3) ◽  
pp. 495-501 ◽  
Author(s):  
H. Sadeghi ◽  
A. L. Lumanglas ◽  
W. R. Baumbach ◽  
B. S. Wang
Keyword(s):  
Growth Hormone ◽  
Monoclonal Antibodies ◽  
Binding Assay ◽  
Interaction Analysis ◽  
Binding Protein ◽  
Antigenic Determinants ◽  
Binding Assays ◽  
Competition Binding Assay ◽  
Competition Binding ◽  
Sequential Binding

ABSTRACT The properties of four independent lines of monoclonal antibodies (MAbs) specific to rat GH-binding protein (GHBP) were examined. Three MAbs, designated GHR-12, GHR-13 and GHR-16, were raised against the entire GHBP molecule. The fourth MAb, designated as GHBP4·3, was raised against the 17 amino acid residues at the C-terminal end of rat GHBP. The interaction of these antibodies with GHBP and their effect on GH binding to GHBP were analysed by conventional competition binding assays and surface plasmon resonance, i.e. with a Biospecific Interaction Analysis (BIAcore) instrument. The binding affinity of these MAbs to GHBP ranged from 29 nmol/l to 30·9 pmol/l. The pair-wise antibody binding to GHBP on BIAcore suggested that GHR-13 and GHR-16 recognized different antigenic determinants while part of the GHR-12 epitope might be shared with the other antibodies. The antibodies inhibited the interaction of GH with GHBP in the competition binding assay. However, in sequential binding on the BIAcore instrument, they were able to bind GHBP after its interaction with GH, indicating that the inhibition observed in the competition binding assay resulted from steric hindrance rather than direct interference with the GH-binding site of GHBP. The present findings, therefore, suggest that these antibodies are useful for investigating GHBP and its interaction with GH. Journal of Endocrinology (1993) 139, 495–501

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