scholarly journals 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

2006 ◽  
Vol 13 (5) ◽  
pp. 594-597 ◽  
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.

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

2010 ◽  
Vol 79 (1) ◽  
pp. 512-517 ◽  
Author(s):  
Yong-Feng Fu ◽  
Meng Feng ◽  
Kenji Ohnishi ◽  
Tamon Kimura ◽  
Johbu Itoh ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Surface Antigen ◽  
Heavy Chain ◽  
Germ Line ◽  
Human Monoclonal Antibody ◽  
Passive Immunization ◽  
Positive Staining ◽  
Immunoglobulin Gene ◽  
Fab Fragment ◽  
Fab Fragments

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.

scholarly journals Functional Activities and Immunoglobulin Variable Regions of Human and Murine Monoclonal Antibodies Specific for the P1.7 PorA Protein Loop of Neisseria meningitidis

2000 ◽  
Vol 68 (4) ◽  
pp. 1871-1878 ◽  
Author(s):  
Jianfu Wang ◽  
Gary A. Jarvis ◽  
Mark Achtman ◽  
Einar Rosenqvist ◽  
Terje E. Michaelsen ◽  
...  
Keyword(s):  
Human Monoclonal Antibody ◽  
Variable Region ◽  
Effector Function ◽  
Enzyme Linked Immunosorbent Assay ◽  
Molecular Characteristics ◽  
Fab Fragment ◽  
Region Gene ◽  
Variable Regions ◽  
Group B ◽  
Murine Mabs

ABSTRACT The meningococcal PorA protein is considered a promising vaccine candidate. Although much is understood regarding the structure of PorA proteins, little is known about the structure-function relationships of PorA antibodies. The aim of this study was to compare the functional and molecular characteristics of a human monoclonal antibody (MAb) and three murine MAbs specific for the PorA P1.7 serosubtype. Murine MAbs 207,B-4 (immunoglobulin G2a [IgG2a]) and MN14C11.6 (IgG2a) were both bactericidal and opsonophagocytic for P1.7-expressing meningococci, whereas human MAb SS269 (IgG3) and murine MAb 208,D-5 (IgA) initiated neither effector function. Epitope mapping with synthetic peptides revealed that MAbs 207,B-4 and 208,D-5 recognized the sequence ASGQ, which is the same specificity motif that a previous study had established for SS269 and MN14C11.6. Nucleotide and amino acid sequence analyses of the variable regions of the four MAbs showed that the SS269 VH region belonged to the VH3 family and was approximately 70% homologous to those of the murine MAbs which were all from the 7183 family, whereas the SS269 VL region belonged to the Vλ1-b family and was less than 40% homologous to those of the murine MAbs which were all members of the Vκ1 family. The Fab fragment of SS269 was cloned and expressed in Escherichia coli and was shown by enzyme-linked immunosorbent assay analyses to bind as well as intact SS269 MAb to P1.7,16 serosubtype group B strain 44/76. We conclude that distinct differences exist in the effector function activities and variable region gene sequences of human and murine P1.7-specific MAbs despite their recognition of similar epitopes.

scholarly journals A Quantitative ELISA Protocol for Detection of Specific Human IgG Against the SARS-CoV-2 Spike Protein

Vaccines ◽  
2021 ◽  
Vol 9 (7) ◽  
pp. 770
Author(s):  
Rémi Vernet ◽  
Emily Charrier ◽  
Julien Grogg ◽  
Nicolas Mach
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Immunosorbent Assay ◽  
Spike Protein ◽  
Quantitative Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibodies ◽  
Human Igg ◽  
Microtiter Plates ◽  
Global Priority ◽  
Humoral Responses

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a worldwide pandemic with at least 3.8 million deaths to date. For that reason, finding an efficient vaccine for this virus quickly became a global priority. The majority of vaccines now marketed are based on the SARS‑CoV‑2 spike protein that has been described as the keystone for optimal immunization. In order to monitor SARS‑CoV‑2 spike-specific humoral responses generated by immunization or infection, we have developed a robust and reproducible enzyme-linked immunosorbent assay (ELISA) protocol. This protocol describes a method for quantitative detection of IgG antibodies against the SARS‑CoV‑2 spike protein using antigen-coated microtiter plates. Results showed that antibodies could be quantified between the range of 1.953 ng/mL to 500 ng/mL with limited inter- and intra-assay variability.

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

2007 ◽  
Vol 75 (7) ◽  
pp. 3614-3620 ◽  
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.

scholarly journals Human Neutralizing Fab Molecules against Severe Acute Respiratory Syndrome Coronavirus Generated by Phage Display

2006 ◽  
Vol 13 (8) ◽  
pp. 953-957 ◽  
Author(s):  
Xiaoping Kang ◽  
Bao-an Yang ◽  
Yuyang Hu ◽  
Hui Zhao ◽  
Wei Xiong ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Severe Acute Respiratory Syndrome ◽  
Dna Sequence ◽  
Heavy Chain ◽  
Cytopathic Effect ◽  
Treated Group ◽  
Spike Protein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Reverse Transcription Pcr ◽  
Neutralizing Capacity

ABSTRACT Human recombinant Fab fragments specific for the spike protein of severe acute respiratory syndrome coronavirus (SARS-CoV) were screened from a human Fab library, which was generated from RNAs from peripheral lymphocytes of convalescent SARS patients. Among 50 randomly picked clones, 12 Fabs specially reacted with S protein by an enzyme-linked immunosorbent assay. The microneutralizing test showed that one clone, designated M1A, had neutralizing activity on Vero E6 cells against SARS-CoV. DNA sequence analysis indicated that the light- and heavy-chain genes of M1A Fab belong to the κ2a and 4f families, respectively. A neutralizing test on purified M1A demonstrated that 0.5 mg/ml of M1A completely inhibited SARS-CoV activity, with an absence of cytopathic effect for 7 days. Real-time fluorescence reverse transcription-PCR also proved the neutralizing capacity of M1A. These data showed that the number of virus copies was significantly reduced in the M1A-treated group, suggesting an important role for M1A in passive immunoprophylaxis against the SARS virus.

scholarly journals Human Milk Antibodies Against SARS-CoV-2: A Longitudinal Follow-Up Study

2021 ◽  
pp. 089033442110301
Author(s):  
Hannah G. Juncker ◽  
M. Romijn ◽  
Veerle N. Loth ◽  
Tom G. Caniels ◽  
Christianne J.M. de Groot ◽  
...  
Keyword(s):  
Human Milk ◽  
Clinical Symptoms ◽  
Immunoglobulin A ◽  
Spike Protein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Follow Up Study ◽  
Lactating Mothers ◽  
Milk Antibodies ◽  
Over Time

Background: Human milk contains antibodies against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) following Coronavirus Disease 2019 (COVID-19). These antibodies may serve as protection against COVID-19 in infants. However, the evolution of these human milk antibodies over time is unclear. Research Aim: To elucidate the evolution of immunoglobulin A (IgA) against SARS-CoV-2 in human milk after a SARS-CoV-2 infection. Methods: This longitudinal follow-up study included lactating mothers ( N = 24) who had participated in the COVID MILK study. To assess the evolution of SARS-CoV-2 antibodies, serum and human milk samples were collected 14–143 days after the onset of clinical symptoms related to COVID-19. Enzyme-Linked ImmunoSorbent Assay was used to detect antibodies against the ectodomain of the SARS-CoV-2 spike protein. Results: SARS-CoV-2 antibodies remain present up to 5 months (143 days) in human milk after onset of COVID-19 symptoms. Overall, SARS-CoV-2 IgA in human milk seems to gradually decrease over time. Conclusion: Human milk from SARS-CoV-2 convalescent lactating mothers contains specific IgA antibodies against SARS-CoV-2 spike protein up to at least 5 months post-infection. Passive viral immunity can be transferred via human milk and may serve as protection for infants against COVID-19. Dutch Trial Register on May 1st, 2020, number: NL 8575, URL: https://www.trialregister.nl/trial/8575 .

scholarly journals Immunological imprinting of the antibody response in COVID-19 patients

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Teresa Aydillo ◽  
Alexander Rombauts ◽  
Daniel Stadlbauer ◽  
Sadaf Aslam ◽  
Gabriela Abelenda-Alonso ◽  
...  
Keyword(s):  
Immune Response ◽  
Severe Acute Respiratory Syndrome ◽  
Antibody Response ◽  
Humoral Immune Response ◽  
Nucleocapsid Protein ◽  
Spike Protein ◽  
Ongoing Research ◽  
Variable Regions ◽  
Humoral Immune ◽  
Human Coronaviruses

AbstractIn addition to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), humans are also susceptible to six other coronaviruses, for which consecutive exposures to antigenically related and divergent seasonal coronaviruses are frequent. Despite the prevalence of COVID-19 pandemic and ongoing research, the nature of the antibody response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is unclear. Here we longitudinally profile the early humoral immune response against SARS-CoV-2 in hospitalized coronavirus disease 2019 (COVID-19) patients and quantify levels of pre-existing immunity to OC43, HKU1 and 229E seasonal coronaviruses, and find a strong back-boosting effect to conserved but not variable regions of OC43 and HKU1 betacoronaviruses spike protein. However, such antibody memory boost to human coronaviruses negatively correlates with the induction of IgG and IgM against SARS-CoV-2 spike and nucleocapsid protein. Our findings thus provide evidence of immunological imprinting by previous seasonal coronavirus infections that can potentially modulate the antibody profile to SARS-CoV-2 infection.

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