Generation of a Neutralizing Human Monoclonal Antibody Fab Fragment to Surface Antigen 1 ofToxoplasma gondiiTachyzoites

ABSTRACTA combinatorial immunoglobulin gene library was constructed from lymphocytes in peripheral blood of a patient with toxoplasmosis and screened for production of human monoclonal antibody Fab fragments to recombinant surface antigen 1 (SAG1) ofToxoplasma gondii. Two Fab clones, Tox203 and Tox1403, which consisted of a common heavy chain and different light chains, showed positive staining on the entire surface of tachyzoites in confocal microscopy. Sequence analysis of the heavy-chain gene revealed that the closest germ line V segments were VH3-23. The germ line D segment was D1-7, and the closest germ line J segment was JH4. In the light-chain genes, the closest germ line V segment was Vκ1-17 with the Jκ1 or Jκ4 segments. The dissociation constants of these Fab fragments with recombinant SAG1 were 3.09 × 10−9M for Tox203 and 2.01 × 10−8M for Tox1403, indicating that the affinity of Tox203 was 7 times higher than that of Tox1403. Preincubation ofT. gondiitachyzoites with Tox203 significantly inhibited their attachment to cultured MDBK cells. Passive immunization of mice with Tox203 also significantly reduced mortality after challenge withT. gondiitachyzoites. This is the first report of bacterial expression of human monoclonal antibody Fab fragments to SAG1 ofT. gondii. These results also demonstrate that human Fab fragments to SAG1 might be applicable for immunoprophylaxis of toxoplasmosis.

Download Full-text

Production of High-Affinity Human Monoclonal Antibody Fab Fragments to the 19-Kilodalton C-Terminal Merozoite Surface Protein 1 of Plasmodium falciparum

Infection and Immunity ◽

10.1128/iai.00062-07 ◽

2007 ◽

Vol 75 (7) ◽

pp. 3614-3620 ◽

Cited By ~ 11

Author(s):

Xun-Jia Cheng ◽

Hitoshi Hayasaka ◽

Katsuomi Watanabe ◽

Yan-Lin Tao ◽

Jin-Ye Liu ◽

...

Keyword(s):

Monoclonal Antibody ◽

Germ Line ◽

Human Monoclonal Antibody ◽

Surface Protein ◽

Merozoite Surface Protein ◽

Immunoglobulin Gene ◽

Gene Library ◽

High Affinity ◽

Merozoite Surface Protein 1 ◽

Fab Fragments

ABSTRACT A combinatorial immunoglobulin gene library was constructed from peripheral blood lymphocytes of eight patients infected with Plasmodium falciparum and was screened for the production of human monoclonal antibody Fab fragments to the C-terminal 19-kDa fragment of P. falciparum merozoite surface protein 1 (MSP-119). Three Fab clones recognized recombinant MSP-119 under nonreducing conditions. Indirect immunofluorescence microscopy demonstrated that three Fab clones stained the surfaces of late trophozoites/schizonts and merozoites of the FCR3 and 3D7 strains, suggesting the Fabs' reactivities to a conserved epitope. Sequence analysis of the heavy-chain genes revealed that the closest germ line V segments were VH1-8 and VH7-81, with 91% to 98% homology. The closest germ line D segment was D3-10, and the closest germ line J segment was JH4 or JH5, with 90% to 97% homology. In the light-chain genes, the closest germ line V segment was A27 for the Jκ2, Jκ4, and Jκ5 segments. The dissociation constants of these Fab fragments for recombinant MSP-119 ranged from 1.09 × 10−9 to 2.66 × 10−9 M. The binding of the three Fab fragments to MSP-119 was competitively inhibited by the anti-MSP-119 mouse monoclonal antibody 12.8, which inhibits erythrocyte invasion by merozoites. However, the human Fab fragment with the highest affinity did not inhibit in vitro growth of P. falciparum. This is the first report of gene analysis and bacterial expression of human monoclonal antibodies to P. falciparum MSP-119. The combinatorial immunoglobulin gene library derived from malaria patients provides a potential tool for producing high-affinity human antibodies specific for P. falciparum.

Download Full-text

Preparation of Recombinant Human Monoclonal Antibody Fab Fragments Specific for Entamoeba histolytica

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.6.3.383-387.1999 ◽

1999 ◽

Vol 6 (3) ◽

pp. 383-387 ◽

Cited By ~ 12

Author(s):

Hiroshi Tachibana ◽

Xun-Jia Cheng ◽

Katsuomi Watanabe ◽

Masataka Takekoshi ◽

Fumiko Maeda ◽

...

Keyword(s):

Escherichia Coli ◽

Monoclonal Antibody ◽

Entamoeba Histolytica ◽

Heavy Chain ◽

Light Chain ◽

Dna Sequences ◽

Human Monoclonal Antibody ◽

Fab Fragment ◽

Fab Fragments ◽

Reference Strains

ABSTRACT Genes coding for human antibody Fab fragments specific forEntamoeba histolytica were cloned and expressed inEscherichia coli. Lymphocytes were separated from the peripheral blood of a patient with an amebic liver abscess. Poly(A)+ RNA was isolated from the lymphocytes, and then genes coding for the light chain and Fd region of the heavy chain were amplified by a reverse transcriptase PCR. The amplified DNA fragments were ligated with a plasmid vector and were introduced intoEscherichia coli. Three thousand colonies were screened for the production of antibodies to E. histolytica HM-1:IMSS by an indirect fluorescence-antibody (IFA) test. Lysates from fiveEscherichia coli clones were positive. Analysis of the DNA sequences of the five clones showed that three of the five heavy-chain sequences and four of the five light-chain sequences differed from each other. When the reactivities of the Escherichia colilysates to nine reference strains of E. histolytica were examined by the IFA test, three Fab fragments with different DNA sequences were found to react with all nine strains and another Fab fragment was found to react with seven strains. None of the four human monoclonal antibody Fab fragments reacted with Entamoeba dispar reference strains or with other enteric protozoan parasites. These results indicate that the bacterial expression system reported here is effective for the production of human monoclonal antibodies specific for E. histolytica. The recombinant human monoclonal antibody Fab fragments may be applicable for distinguishing E. histolytica from E. dispar and for use in the serodiagnosis of amebiasis.

Download Full-text

Immunoglobulin gene rearrangements in normal mouse B cells

Molecular and Cellular Biology ◽

10.1128/mcb.2.7.829-836.1982 ◽

1982 ◽

Vol 2 (7) ◽

pp. 829-836

Author(s):

P Early ◽

C Nottenburg ◽

I Weissman ◽

L Hood

Keyword(s):

B Cells ◽

Heavy Chain ◽

Germ Line ◽

Gene Segment ◽

Immunoglobulin M ◽

Chain Gene ◽

Allelic Exclusion ◽

Immunoglobulin Gene ◽

Spleen Cells ◽

Surface Immunoglobulin

We have analyzed the structure of rearranged mu heavy-chain genes obtained from the genomic DNA of normal BALB/c mouse spleen cells expressing surface immunoglobulin M. Examples were found of two types of nonproductive rearrangements, which may be responsible for allelic exclusion in normal B cells. In one of these rearrangements, a germ line D gene segment has joined to the JH4 gene segment but no V/D joining has occurred. We present evidence that D gene segments lie as a cluster between V and J gene segments in the germ line. A comparison of conserved sequences in V and D gene segments suggests that the D gene segments, which are found only in the heavy-chain gene family, may have evolved from V gene segments similar to the Vk family.

Download Full-text

Structure of a human monoclonal antibody Fab fragment against gp41 of human immunodeficiency virus type 1.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.89.15.7154 ◽

1992 ◽

Vol 89 (15) ◽

pp. 7154-7158 ◽

Cited By ~ 40

Author(s):

X. M. He ◽

F. Ruker ◽

E. Casale ◽

D. C. Carter

Keyword(s):

Monoclonal Antibody ◽

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Human Monoclonal Antibody ◽

Fab Fragment ◽

Immunodeficiency Virus

Download Full-text

Production and Modification of Human Monoclonal Antibody Fab Fragments to the 19-Kilodalton C-Terminal Merozoite Surface Protein 1 of Plasmodium falciparum

International Journal of Infectious Diseases ◽

10.1016/j.ijid.2008.05.835 ◽

2008 ◽

Vol 12 ◽

pp. e311-e312

Author(s):

H. Tachibana ◽

X. Cheng ◽

Y. Tao ◽

Y. Fu ◽

E. Yoshihara ◽

...

Keyword(s):

Monoclonal Antibody ◽

Plasmodium Falciparum ◽

Human Monoclonal Antibody ◽

Surface Protein ◽

Merozoite Surface Protein ◽

Merozoite Surface Protein 1 ◽

Fab Fragments

Download Full-text

A Human Monoclonal Antibody against Hepatitis B Surface Antigen with Potent Neutralizing Activity

PLoS ONE ◽

10.1371/journal.pone.0125704 ◽

2015 ◽

Vol 10 (4) ◽

pp. e0125704 ◽

Cited By ~ 16

Author(s):

Antonella Cerino ◽

Corinna M. Bremer ◽

Dieter Glebe ◽

Mario U. Mondelli

Keyword(s):

Monoclonal Antibody ◽

Hepatitis B ◽

Surface Antigen ◽

Hepatitis B Surface Antigen ◽

Human Monoclonal Antibody ◽

Neutralizing Activity

Download Full-text

Production of an Anti-Severe Acute Respiratory Syndrome (SARS) Coronavirus Human Monoclonal Antibody Fab Fragment by Using a Combinatorial Immunoglobulin Gene Library Derived from Patients Who Recovered from SARS

Clinical and Vaccine Immunology ◽

10.1128/cvi.13.5.594-597.2006 ◽

2006 ◽

Vol 13 (5) ◽

pp. 594-597 ◽

Cited By ~ 8

Author(s):

Jinye Liu ◽

Hongxia Shao ◽

Yanlin Tao ◽

Bin Yang ◽

Lisheng Qian ◽

...

Keyword(s):

Severe Acute Respiratory Syndrome ◽

Human Monoclonal Antibody ◽

Antibody Fragments ◽

Spike Protein ◽

Enzyme Linked Immunosorbent Assay ◽

Immunoglobulin Gene ◽

Gene Library ◽

Fab Fragment ◽

Human Antibodies ◽

Infected Cells

ABSTRACT A combinatorial human immunoglobulin gene library was constructed from the peripheral lymphocytes of two patients who recovered from severe acute respiratory syndrome (SARS). The library was screened for the production of Fab antibody fragments to a recombinant spike protein of SARS-associated coronavirus (SARS-CoV). One Fab clone, AS3-3, reacted with the spike protein in an enzyme-linked immunosorbent assay. The dissociation constant of AS3-3 was 1.98 × 10−8 M. Immunofluorescent microscopy revealed that it reacted with SARS-CoV-infected cells. The library seems to be a potent tool for the production of human antibodies to SARS-CoV.

Download Full-text