scholarly journals Cryptosporidium parvum Sporozoite Pellicle Antigen Recognized by a Neutralizing Monoclonal Antibody Is a β-Mannosylated Glycolipid

1999 ◽  
Vol 67 (3) ◽  
pp. 1317-1322 ◽  
Author(s):  
Michael W. Riggs ◽  
Michael R. McNeil ◽  
Lance E. Perryman ◽  
Alice L. Stone ◽  
Michael S. Scherman ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Silicic Acid ◽  
Cryptosporidium Parvum ◽  
Protozoan Parasite ◽  
Subcellular Location ◽  
Protective Antigens ◽  
Silicic Acid Chromatography ◽  
Target Epitope ◽  
Neutralizing Monoclonal Antibody ◽  
Surface Localization

ABSTRACT The protozoan parasite Cryptosporidium parvum is an important cause of diarrhea in humans, calves, and other mammals worldwide. No approved vaccines or parasite-specific drugs are currently available for the control of cryptosporidiosis. To effectively immunize against C. parvum, identification and characterization of protective antigens are required. We previously identified CPS-500, a conserved, neutralization-sensitive antigen ofC. parvum sporozoites and merozoites defined by monoclonal antibody 18.44. In the present study, the biochemical characteristics and subcellular location of CPS-500 were determined. CPS-500 was chloroform extractable and eluted with acetone and methanol in silicic acid chromatography, consistent with being a polar glycolipid. Following chloroform extraction and silicic acid chromatography, CPS-500 was isolated by high-pressure liquid chromatography for glycosyl analysis, which indicated the presence of mannose and inositol. To identify which component of CPS-500 comprised the neutralization-sensitive epitope recognized by 18.44, the ability of the monoclonal antibody to bind CPS-500 treated with proteases, or with α- or β-glycosidases, was determined. Monoclonal antibody 18.44 did not bind antigen treated with β-d-mannosidase but did bind antigen treated with α-d-mannosidase, other α- or β-glycosidases, or a panel of proteases. These data indicated that the target epitope was dependent on terminal β-d-mannopyranosyl residues. By immunoelectron microscopy, 18.44 binding was localized to the pellicle and an intracytoplasmic tubulovesicular network in sporozoites. Monoclonal antibody 18.44 also bound to antigen deposited and released onto substrate over the course travelled by gliding sporozoites and merozoites. Surface localization, adhesion and release during locomotion, and neutralization sensitivity suggest that CPS-500 may be involved in motility and invasion processes of the infective zoite stages.

scholarly journals High Diversity of Cryptosporidium Species and Subtypes Identified in Cryptosporidiosis Acquired in Sweden and Abroad

Pathogens ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 523
Author(s):  
Marianne Lebbad ◽  
Jadwiga Winiecka-Krusnell ◽  
Christen Rune Stensvold ◽  
Jessica Beser
Keyword(s):  
Cryptosporidium Parvum ◽  
Ribosomal Rna ◽  
Diarrheal Disease ◽  
Protozoan Parasite ◽  
Small Subunit ◽  
Cryptosporidium Species ◽  
Intestinal Protozoan ◽  
Genotype I ◽  
Glycoprotein Gene ◽  
High Diversity

The intestinal protozoan parasite Cryptosporidium is an important cause of diarrheal disease worldwide. The aim of this study was to expand the knowledge on the molecular epidemiology of human cryptosporidiosis in Sweden to better understand transmission patterns and potential zoonotic sources. Cryptosporidium-positive fecal samples were collected between January 2013 and December 2014 from 12 regional clinical microbiology laboratories in Sweden. Species and subtype determination was achieved using small subunit ribosomal RNA and 60 kDa glycoprotein gene analysis. Samples were available for 398 patients, of whom 250 (63%) and 138 (35%) had acquired the infection in Sweden and abroad, respectively. Species identification was successful for 95% (379/398) of the samples, revealing 12 species/genotypes: Cryptosporidium parvum (n = 299), C. hominis (n = 49), C. meleagridis (n = 8), C. cuniculus (n = 5), Cryptosporidium chipmunk genotype I (n = 5), C. felis (n = 4), C. erinacei (n = 2), C. ubiquitum (n = 2), and one each of C. suis, C. viatorum, C. ditrichi, and Cryptosporidium horse genotype. One patient was co-infected with C. parvum and C. hominis. Subtyping was successful for all species/genotypes, except for C. ditrichi, and revealed large diversity, with 29 subtype families (including 4 novel ones: C. parvum IIr, IIs, IIt, and Cryptosporidium horse genotype VIc) and 81 different subtypes. The most common subtype families were IIa (n = 164) and IId (n = 118) for C. parvum and Ib (n = 26) and Ia (n = 12) for C. hominis. Infections caused by the zoonotic C. parvum subtype families IIa and IId dominated both in patients infected in Sweden and abroad, while most C. hominis cases were travel-related. Infections caused by non-hominis and non-parvum species were quite common (8%) and equally represented in cases infected in Sweden and abroad.

A Factor VIII Neutralizing Monoclonal Antibody and a Human Inhibitor Alloantibody Recognizing Epitopes in the C2 Domain Inhibit Factor VIII Binding to von Willebrand Factor and to Phosphatidylserine

1993 ◽  
Vol 69 (03) ◽  
pp. 240-246 ◽  
Author(s):  
Midori Shima ◽  
Dorothea Scandella ◽  
Akira Yoshioka ◽  
Hiroaki Nakai ◽  
Ichiro Tanaka ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Amino Acid ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Light Chain ◽  
Amino Acid Residues ◽  
Amino Terminal ◽  
Neutralizing Monoclonal Antibody ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryA neutralizing monoclonal antibody, NMC-VIII/5, recognizing the 72 kDa thrombin-proteolytic fragment of factor VIII light chain was obtained. Binding of the antibody to immobilized factor VIII (FVIII) was completely blocked by a light chain-specific human alloantibody, TK, which inhibits FVIII activity. Immunoblotting analysis with a panel of recombinant protein fragments of the C2 domain deleted from the amino-terminal or the carboxy-terminal ends demonstrated binding of NMC-VIII/5 to an epitope located between amino acid residues 2170 and 2327. On the other hand, the epitope of the inhibitor alloantibody, TK, was localized to 64 amino acid residues from 2248 to 2312 using the same recombinant fragments. NMC-VIII/5 and TK inhibited FVIII binding to immobilized von Willebrand factor (vWF). The IC50 of NMC-VIII/5 for the inhibition of binding to vWF was 0.23 μg/ml for IgG and 0.2 μg/ml for F(ab)'2. This concentration was 100-fold lower than that of a monoclonal antibody NMC-VIII/10 which recognizes the amino acid residues 1675 to 1684 within the amino-terminal portion of the light chain. The IC50 of TK was 11 μg/ml by IgG and 6.3 μg/ml by F(ab)'2. Furthermore, NMC-VIII/5 and TK also inhibited FVIII binding to immobilized phosphatidylserine. The IC50 for inhibition of phospholipid binding of NMC-VIII/5 and TK (anti-FVIII inhibitor titer of 300 Bethesda units/mg of IgG) was 10 μg/ml.

scholarly journals Development of a New Highly Selective Monoclonal Antibody against Preferentially Expressed Antigen in Melanoma (PRAME) and Identification of the Target Epitope by Bio-Layer Interferometry

2021 ◽  
Vol 22 (6) ◽  
pp. 3166
Author(s):  
Jwala Priyadarsini Sivaccumar ◽  
Antonio Leonardi ◽  
Emanuela Iaccarino ◽  
Giusy Corvino ◽  
Luca Sanguigno ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Western Blotting ◽  
Cancer Biomarkers ◽  
Trypsin Digestion ◽  
Protein Fragments ◽  
Target Epitope ◽  
Potential Activity ◽  
Antibody Epitopes

Background: Monoclonal antibodies (mAbs) against cancer biomarkers are key reagents in diagnosis and therapy. One such relevant biomarker is a preferentially expressed antigen in melanoma (PRAME) that is selectively expressed in many tumors. Knowing mAb’s epitope is of utmost importance for understanding the potential activity and therapeutic prospective of the reagents. Methods: We generated a mAb against PRAME immunizing mice with PRAME fragment 161–415; the affinity of the antibody for the protein was evaluated by ELISA and SPR, and its ability to detect the protein in cells was probed by cytofluorimetry and Western blotting experiments. The antibody epitope was identified immobilizing the mAb on bio-layer interferometry (BLI) sensor chip, capturing protein fragments obtained following trypsin digestion and performing mass spectrometry analyses. Results: A mAb against PRAME with an affinity of 35 pM was obtained and characterized. Its epitope on PRAME was localized on residues 202–212, taking advantage of the low volumes and lack of fluidics underlying the BLI settings. Conclusions: The new anti-PRAME mAb recognizes the folded protein on the surface of cell membranes suggesting that the antibody’s epitope is well exposed. BLI sensor chips can be used to identify antibody epitopes.

scholarly journals Structural elements of primary CCR5-using HIV-1 gp120 proteins influencing sensitivity and resistance to the broadly neutralizing monoclonal antibody b12

Virology ◽  
2012 ◽  
Vol 432 (2) ◽  
pp. 394-404
Author(s):  
Jasminka Sterjovski ◽  
Melissa J. Churchill ◽  
Anne Ellett ◽  
Steve L. Wesselingh ◽  
Paul A. Ramsland ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Structural Elements ◽  
Neutralizing Monoclonal Antibody ◽  
Hiv 1

scholarly journals Structure of adeno-associated virus-2 in complex with neutralizing monoclonal antibody A20

Virology ◽  
2012 ◽  
Vol 431 (1-2) ◽  
pp. 40-49 ◽  
Author(s):  
Dustin M. McCraw ◽  
Jason K. O’Donnell ◽  
Kenneth A. Taylor ◽  
Scott M. Stagg ◽  
Michael S. Chapman
Keyword(s):  
Monoclonal Antibody ◽  
Adeno Associated Virus ◽  
Neutralizing Monoclonal Antibody

The RAV12 Monoclonal Antibody Recognizes the N-Linked Glycotope RAAG12: Expression in Human Normal and Tumor Tissues

2009 ◽  
Vol 133 (9) ◽  
pp. 1403-1412
Author(s):  
Suzanne K. Coberly ◽  
Francine Z. Chen ◽  
Mark P. Armanini ◽  
Yan Chen ◽  
Peter F. Young ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Antigen Expression ◽  
Subcellular Location ◽  
Carbohydrate Antigen ◽  
Cytoplasmic Staining ◽  
Safety Issues ◽  
Normal Tissues ◽  
Tumor Tissues ◽  
Associated Proteins

Abstract Context.—RAAG12 is a primate-restricted N-linked carbohydrate antigen present on multiple membrane-associated proteins. RAAG12 is recognized by the RAV12 monoclonal antibody. RAV12 binds to RAAG12-expressing gastrointestinal adenocarcinomas, modifies growth factor-mediated signaling, induces oncotic cell death in vitro, and has antitumor activity toward gastrointestinal tumor xenografts. Objective.—To determine the expression pattern of RAAG12 in normal and tumor tissue to identify indications for clinical study and potential safety issues. Design.—Immunohistochemistry of 36 normal human tissues and a broad range of tumor tissues to profile RAAG12 expression. Results.—More than 90% of colon, gastric, and pancreatic adenocarcinomas expressed RAAG12, and expression was uniform in most samples. Expression of RAAG12 at lower frequency and/or uniformity was observed in other cancers, including esophageal, ovarian, liver, breast, and prostate carcinomas and adenocarcinomas. Similar RAAG12 expression was observed between primary and metastatic colon adenocarcinomas. No staining was seen on cardiovascular, endocrine, neuromuscular, hematopoietic, or nervous system tissue from non–tumor-bearing individuals. RAAG12 was expressed on mucosal and glandular/ductal epithelium. The gastrointestinal tract mucosa and pancreatic/biliary ducts displayed the most uniform reactivity. RAAG12 exhibited differential subcellular localization in these normal, compared with tumor, tissues. Normal polarized epithelia primarily displayed apical membrane and cytoplasmic staining, whereas tumors exhibited whole membrane staining that increased with decreasing differentiation. Conclusions.—High expression of RAAG12 on tumors of gastrointestinal origin suggests these cancers are appropriate targets for RAV12 therapy. Differential subcellular location of RAAG12 on normal epithelia may limit accessibility of RAV12 to the subset of normal tissues that exhibit antigen expression.

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