Antibody production in goat milk serum after virus instillation of goat mammary gland. VII. Biochemical studies on sham infection with rabies and other non-propagating agents

1975 ◽  
Vol 21 (5) ◽  
pp. 655-661 ◽  
Author(s):  
Arthur E. Pasieka ◽  
L. F. Guerin ◽  
Chas. A. Mitchell
Keyword(s):  
Mammary Gland ◽  
Rabies Virus ◽  
Neutralizing Antibodies ◽  
Analytical Ultracentrifugation ◽  
Sedimentation Coefficient ◽  
Goat Milk ◽  
Neutralizing Antibody ◽  
Antibody Activity ◽  
Gel Chromatography ◽  
Milk Serum

This report describes the development of a method for the production and isolation of neutralizing antibodies against rabies virus (ERA strain) and other agents in the goat mammary gland during active lactation. The rabies virus did not propagate in the gland, but neutralizing antibodies were produced by serial instillations of the antigen. This process was termed 'sham infection.' Antibodies first appeared in the milk about the 20th day and the titer increased until the 28th day. The antibodies were found to be associated with the lactogammaglobulin. The milk globulins were fractionated by gel chromatography and the fraction containing the antibody was isolated. This fraction was further purified and characterized by immunodiffusion, immunoelectrophoresis, and an analytical ultracentrifugation technique. The neutralizing antibody activity after the 29th day was associated with the milk serum IgG fraction. This active fraction was found to have a sedimentation coefficient of 6.8 Svedberg units.

Antibody production in milk serum of goats experimentally infected with Neisseria gonorrhoeae

1976 ◽  
Vol 22 (8) ◽  
pp. 1113-1119 ◽  
Author(s):  
A. E. Pasieka ◽  
F. E. Ashton ◽  
R. Wallace ◽  
F. Ota ◽  
A. Ryan ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Antibody Response ◽  
Neisseria Gonorrhoeae ◽  
Analytical Ultracentrifugation ◽  
Gel Filtration ◽  
Goat Milk ◽  
Purification And Characterization ◽  
Colony Type ◽  
Milk Serum

Instillation of the goat's mammary gland with Neisseria gonorrhoeae colony type Tl has elicited an antibody response in the goat milk serum (GMS). Purification, and characterization of the GMS by gel filtration, electrophoresis, immunodiffusion, analytical ultracentrifugation. And serological analyses demonstrated that the active immune component was mainly in the IgA and IgG fractions (F2 and F3) of GMS.

scholarly journals Characterization and Implementation of a Diverse Simian Immunodeficiency Virus SIVsm Envelope Panel in the Assessment of Neutralizing Antibody Breadth Elicited in Rhesus Macaques by Multimodal Vaccines Expressing the SIVmac239 Envelope

2015 ◽  
Vol 89 (16) ◽  
pp. 8130-8151 ◽  
Author(s):  
Katie M. Kilgore ◽  
Megan K. Murphy ◽  
Samantha L. Burton ◽  
Katherine S. Wetzel ◽  
S. Abigail Smith ◽  
...  
Keyword(s):  
Simian Immunodeficiency Virus ◽  
Nonhuman Primate ◽  
Neutralizing Antibodies ◽  
Rhesus Macaques ◽  
Neutralizing Antibody ◽  
Cd40 Ligand ◽  
Antibody Activity ◽  
Immunodeficiency Virus ◽  
Antibody Profile ◽  
Hiv 1

ABSTRACTAntibodies that can neutralize diverse viral strains are likely to be an important component of a protective human immunodeficiency virus type 1 (HIV-1) vaccine. To this end, preclinical simian immunodeficiency virus (SIV)-based nonhuman primate immunization regimens have been designed to evaluate and enhance antibody-mediated protection. However, these trials often rely on a limited selection of SIV strains with extreme neutralization phenotypes to assess vaccine-elicited antibody activity. To mirror the viral panels used to assess HIV-1 antibody breadth, we created and characterized a novel panel of 14 genetically and phenotypically diverse SIVsm envelope (Env) glycoproteins. To assess the utility of this panel, we characterized the neutralizing activity elicited by four SIVmac239 envelope-expressing DNA/modified vaccinia virus Ankara vector- and protein-based vaccination regimens that included the immunomodulatory adjuvants granulocyte-macrophage colony-stimulating factor, Toll-like receptor (TLR) ligands, and CD40 ligand. The SIVsm Env panel exhibited a spectrum of neutralization sensitivity to SIV-infected plasma pools and monoclonal antibodies, allowing categorization into three tiers. Pooled sera from 91 rhesus macaques immunized in the four trials consistently neutralized only the highly sensitive tier 1a SIVsm Envs, regardless of the immunization regimen. The inability of vaccine-mediated antibodies to neutralize the moderately resistant tier 1b and tier 2 SIVsm Envs defined here suggests that those antibodies were directed toward epitopes that are not accessible on most SIVsm Envs. To achieve a broader and more effective neutralization profile in preclinical vaccine studies that is relevant to known features of HIV-1 neutralization, more emphasis should be placed on optimizing the Env immunogen, as the neutralization profile achieved by the addition of adjuvants does not appear to supersede the neutralizing antibody profile determined by the immunogen.IMPORTANCEMany in the HIV/AIDS vaccine field believe that the ability to elicit broadly neutralizing antibodies capable of blocking genetically diverse HIV-1 variants is a critical component of a protective vaccine. Various SIV-based nonhuman primate vaccine studies have investigated ways to improve antibody-mediated protection against a heterologous SIV challenge, including administering adjuvants that might stimulate a greater neutralization breadth. Using a novel SIV neutralization panel and samples from four rhesus macaque vaccine trials designed for cross comparison, we show that different regimens expressing the same SIV envelope immunogen consistently elicit antibodies that neutralize only the very sensitive tier 1a SIV variants. The results argue that the neutralizing antibody profile elicited by a vaccine is primarily determined by the envelope immunogen and is not substantially broadened by including adjuvants, resulting in the conclusion that the envelope immunogen itself should be the primary consideration in efforts to elicit antibodies with greater neutralization breadth.

scholarly journals The vaccinia-based Sementis Copenhagen Vector COVID-19 vaccine induces broad and durable cellular and humoral immune responses

2021 ◽  
Author(s):  
Preethi Eldi ◽  
Tamara H Cooper ◽  
Natalie A Prow ◽  
Liang Liu ◽  
Gary K Heinemann ◽  
...  
Keyword(s):  
Immune Responses ◽  
Neutralizing Antibodies ◽  
First Generation ◽  
Neutralizing Antibody ◽  
Immune Memory ◽  
Antibody Activity ◽  
Post Vaccination ◽  
Humoral Immune ◽  
Humoral Immune Responses ◽  
Heterologous Boost

The ongoing COVID-19 pandemic perpetuated by SARS-CoV-2 variants, has highlighted the continued need for broadly protective vaccines that elicit robust and durable protection. Here, the vaccinia virus-based, replication-defective Sementis Copenhagen Vector (SCV) was used to develop a first-generation COVID-19 vaccine encoding the spike glycoprotein (SCV-S). Vaccination of mice rapidly induced polyfunctional CD8 T cells with cytotoxic activity and robust Th1-biased, spike-specific neutralizing antibodies, which are significantly increased following a second vaccination, and contained neutralizing activity against the alpha and beta variants of concern. Longitudinal studies indicated neutralizing antibody activity was maintained up to 9 months post-vaccination in both young and aging mice, with durable immune memory evident even in the presence of pre-existing vector immunity. This immunogenicity profile suggests a potential to expand protection generated by current vaccines in a heterologous boost format, and presents a solid basis for second-generation SCV-based COVID-19 vaccine candidates incorporating additional SARS-CoV-2 immunogens.

ANTIBODY PRODUCTION IN MILK SERUM AFTER VIRUS INSTILLATION OF GOAT MAMMARY GLAND: II. BIOCHEMICAL ISOLATION AND PURIFICATION OF ANTIBODY TO INFLUENZA VIRUS

1967 ◽  
Vol 13 (9) ◽  
pp. 1195-1201 ◽  
Author(s):  
Arthur E. Pasieka ◽  
L. F. Guerin ◽  
Chas. A. Mitchell
Keyword(s):  
Gel Filtration ◽  
Goat Milk ◽  
Protein Band ◽  
Antibody Activity ◽  
Isolation And Purification ◽  
Fraction Collector ◽  
Milk Serum ◽  
Before And After ◽  
The Right ◽  
Working Procedure

Goat milk was obtained from the right lactiferous sinus following instillation of PR8 influenza type A virus. The proteins precipitated from goat milk serum by (NH4)2SO4 were found to contain antibody activity and further purification was undertaken. In the development of the method the protein solutions were subjected to paper chromatography and paper and polyacetate high voltage electrophoresis, both before and after a number of isolation and purification steps. In addition, preparations from each step were run on the Technicon amino acid analyzer for the presence of amino acids and (or) peptides.As a working procedure, a preparation obtained after cold acetone precipitation was subjected to rivanol treatment, and after electrophoresis a single protein band containing the antibody was obtained. This material was further purified by gel filtration using a Bio-Rad P150 column. The antibody containing protein was finally isolated in the eluate by a fraction collector using phosphate buffer as the eluting fluid. There was excellent recovery of the antibody activity in each preparation.

scholarly journals HIV-1 Antibody Neutralization Breadth Is Associated with Enhanced HIV-Specific CD4+T Cell Responses

2015 ◽  
Vol 90 (5) ◽  
pp. 2208-2220 ◽  
Author(s):  
Srinika Ranasinghe ◽  
Damien Z. Soghoian ◽  
Madelene Lindqvist ◽  
Musie Ghebremichael ◽  
Faith Donaghey ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Hiv Infection ◽  
Neutralizing Antibodies ◽  
T Cell Responses ◽  
Neutralizing Antibody ◽  
Antibody Activity ◽  
Cell Responses

ABSTRACTAntigen-specific CD4+T helper cell responses have long been recognized to be a critical component of effective vaccine immunity. CD4+T cells are necessary to generate and maintain humoral immune responses by providing help to antigen-specific B cells for the production of antibodies. In HIV infection, CD4+T cells are thought to be necessary for the induction of Env-specific broadly neutralizing antibodies. However, few studies have investigated the role of HIV-specific CD4+T cells in association with HIV neutralizing antibody activity in vaccination or natural infection settings. Here, we conducted a comprehensive analysis of HIV-specific CD4+T cell responses in a cohort of 34 untreated HIV-infected controllers matched for viral load, with and without neutralizing antibody breadth to a panel of viral strains. Our results show that the breadth and magnitude of Gag-specific CD4+T cell responses were significantly higher in individuals with neutralizing antibodies than in those without neutralizing antibodies. The breadth of Gag-specific CD4+T cell responses was positively correlated with the breadth of neutralizing antibody activity. Furthermore, the breadth and magnitude of gp41-specific, but not gp120-specific, CD4+T cell responses were significantly elevated in individuals with neutralizing antibodies. Together, these data suggest that robust Gag-specific CD4+T cells and, to a lesser extent, gp41-specific CD4+T cells may provide important intermolecular help to Env-specific B cells that promote the generation or maintenance of Env-specific neutralizing antibodies.IMPORTANCEOne of the earliest discoveries related to CD4+T cell function was their provision of help to B cells in the development of antibody responses. Yet little is known about the role of CD4+T helper responses in the setting of HIV infection, and no studies to date have evaluated the impact of HIV-specific CD4+T cells on the generation of antibodies that can neutralize multiple different strains of HIV. Here, we addressed this question by analyzing HIV-specific CD4+T cell responses in untreated HIV-infected persons with and without neutralizing antibodies. Our results indicate that HIV-infected persons with neutralizing antibodies have significantly more robust CD4+T cell responses targeting Gag and gp41 proteins than individuals who lack neutralizing antibodies. These associations suggest that Gag- and gp41-specific CD4+T cell responses may provide robust help to B cells for the generation or maintenance of neutralizing antibodies in natural HIV-infection.

scholarly journals Calculating rabies virus neutralizing antibodies titres by flow cytometry

2002 ◽  
Vol 44 (3) ◽  
pp. 151-154 ◽  
Author(s):  
Juliano BORDIGNON ◽  
Fabiano COMIN ◽  
Sílvia Córdoba P. FERREIRA ◽  
Graciane M. M. CAPORALE ◽  
José Hermênio Cavalcante LIMA FILHO ◽  
...  
Keyword(s):  
Cell Culture ◽  
Flow Cytometry ◽  
Rabies Virus ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Standard Test ◽  
Serum Samples ◽  
Reference Serum ◽  
Infection Inhibition

The determination of the rabies neutralizing antibody (VNA) response after immunization against rabies is an acceptable index of the efficacy of a vaccine and a successful treatment. Several tests have been developed in attempt to improve the assessment of VNA, from mice inoculation to cell-culture fluorescence inhibition tests. All of them, however, present special difficulties in terms of reading or accuracy. The present study describes a neutralization test performed in cell-culture appraised by flow cytometry (FC). Serial dilutions of the serum samples were mixed in vitro with rabies virus before the addition of BHK-21 cells. After 24h-incubation, cells were released by trypsin treatment, fixed and permeabilized with a p-formaldehyde solution and stained with a rabies virus nucleocapsid protein-specific antibody conjugate. The percentage of virus infection inhibition caused by specific antibodies present in the serum were evaluated in a Beckton & Dickinson FACSCalibur® flow cytometer. A correlation curve between the IU/ml content and the percentage of infective inhibition was built with a reference serum and the VNA titers of serum samples were obtained by extrapolation. Titers obtained by FC and standard test showed an effective pairing results (p < 0.01), with a correlation coefficient (r) = 0.7. These results permit to envisage the FC as a suitable technique to evaluate VNA in sera from immunized animals and likely in human serum samples. Nevertheless, new studies comparing FC to gold-standard techniques are required for determining the FC values of Sensibility and Specificity .

scholarly journals The Anamnestic Neutralizing Antibody Response Is Critical for Protection of Mice from Challenge following Vaccination with a Plasmid Encoding the Japanese Encephalitis Virus Premembrane and Envelope Genes

1999 ◽  
Vol 73 (7) ◽  
pp. 5527-5534 ◽  
Author(s):  
Eiji Konishi ◽  
Masaoki Yamaoka ◽  
Khin-Sane-Win ◽  
Ichiro Kurane ◽  
Kazuo Takada ◽  
...  
Keyword(s):  
Japanese Encephalitis ◽  
Neutralizing Antibodies ◽  
Cytotoxic T Lymphocyte ◽  
Neutralizing Antibody ◽  
Immunoglobulin M ◽  
Antibody Activity ◽  
Lethal Challenge ◽  
Challenge Virus ◽  
Coding Regions

ABSTRACT For Japanese encephalitis (JE), we previously reported that recombinant vaccine-induced protection from disease does not prevent challenge virus replication in mice. Moreover, DNA vaccines for JE can provide protection from high challenge doses in the absence of detectable prechallenge neutralizing antibodies. In the present study, we evaluated the role of postchallenge immune responses in determining the outcome of JE virus infection, using mice immunized with a plasmid, pcDNA3JEME, encoding the JE virus premembrane (prM) and envelope (E) coding regions. In the first experiment, 10 mice were vaccinated once (five animals) or twice (remainder) with 100 μg of pcDNA3JEME. All of these mice showed low (6 of 10) or undetectable (4 of 10) levels of neutralizing antibodies. Interestingly, eight of these animals showed a rapid rise in neutralizing antibody following challenge with 10,000 50% lethal doses of JE virus and survived for 21 days, whereas only one of the two remaining animals survived. No unimmunized animals exhibited a rise of neutralizing antibody or survived challenge. Levels of JE virus-specific immunoglobulin M class antibodies were elevated following challenge in half of the unimmunized mice and in the single pcDNA3JEME-immunized mouse that died. In the second experiment, JE virus-specific primary cytotoxic T-lymphocyte (CTL) activity was detected in BALB/c mice immunized once with 100 μg of pcDNA3JEME 4 days after challenge, indicating a strong postchallenge recall of CTLs. In the third experiment, evaluation of induction of CTLs and antibody activity by plasmids containing portions of the prM/E cassette demonstrated that induction of CTL responses alone were not sufficient to prevent death. Finally, we showed that antibody obtained from pcDNA3JEME-immunized mice 4 days following challenge could partially protect recipient mice from lethal challenge. Taken together, these results indicate that neutralizing antibody produced following challenge provides the critical protective component in pcDNA3JEME-vaccinated mice.

scholarly journals Can individuals with low antibody responses to vaccines against other viruses acquire adequate SARS-CoV-2 antibody after vaccination with the BNT162b2 mRNA vaccine?

2021 ◽  
Author(s):  
Wataru Ogura ◽  
Kouki Ohtsuka ◽  
Sachiko Matsuura ◽  
Takahiro Okuyama ◽  
Satsuki Matsushima ◽  
...  
Keyword(s):  
Cohort Study ◽  
Neutralizing Antibodies ◽  
Healthcare Workers ◽  
Neutralizing Antibody ◽  
Antibody Responses ◽  
Antibody Activity ◽  
Low Responders ◽  
Before And After ◽  
Positive Threshold ◽  
Antibody Levels

Objective In Japan, healthcare workers (HCWs) are vaccinated against coronavirus disease (COVID-19) and other contagious viruses (measles, rubella, chickenpox, mumps, and hepatitis B) to prevent nosocomial infection. However, some do not produce sufficient antibodies after vaccination (low responders). This study investigated changes in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody levels among HCWs after SARS-CoV-2 vaccination and assessed whether low responders produced adequate SARS-CoV-2 anti-spike and neutralizing antibodies. Methods We conducted a prospective cohort study of HCWs before and after vaccination with the BNT162b2 mRNA vaccine in a hospital in Tokyo, Japan. The HCWs received two doses of BNT162b2 vaccine, 3 weeks apart. Those whose antibody levels against previous antiviral vaccines did not reach protective antibody levels after receiving two doses were defined as low responders, whereas those who produced adequate antibodies were defined as normal responders. SARS-CoV-2 anti-spike antibodies were measured 11 times from before the first BNT162b2 vaccination to 5 months after the second vaccination. SARS-CoV-2 neutralizing antibody activity was measured twice in low responders, 1 week to 1 month and 5 months after the second vaccination. Results Fifty HCWs were included in the analytic cohort. After vaccination, SARS-CoV-2 anti-spike antibody was detectable in the samples from both responders at each timepoint, but the level was lower at 5 months than at 1 week after the second vaccination. Low responders had SARS-CoV-2 neutralizing antibody activity 1 week to 1 month after the second vaccination, which exceeded the positive threshold after 5 months. Conclusion After BNT162b2 vaccination, low responders acquired adequate SARS-CoV-2 anti-spike and SARS-CoV-2 neutralizing antibodies to prevent SARS-CoV-2. However, SARS-CoV-2 anti-spike antibody levels were lower at 5 months than at 1 week after the second dose of BNT162b2 vaccine in low and normal responders. Therefore, low responders should also receive a third dose of BNT162b2 vaccine.

Antibody production in goat milk serum after virus instillation of goat mammary gland. V. Biochemical isolation and further characterization of antibodies to various influenza and mumps viruses

1970 ◽  
Vol 16 (12) ◽  
pp. 1153-1159 ◽  
Author(s):  
Arthur E. Pasieka ◽  
L. F. Guerin ◽  
Chas. A. Mitchell
Keyword(s):  
Mammary Gland ◽  
Laboratory Animal ◽  
Goat Milk ◽  
Laboratory Animals ◽  
Lactation Period ◽  
Milk Serum ◽  
Therapeutic Uses ◽  
Biochemical Fractionation ◽  
Specific Inhibitors

This report describes the development of a method for the isolation of antibodies produced in the mammary gland and found in the milk after the instillation and propagation of various myxoviruses. Biochemical fractionation and isolation procedures have been modified and improved over our previous initial reports. The antibodies of the various influenza and mumps viruses that propagated in the gland were found to be associated with lactogammaglobulin. This report also demonstrates that the antibodies produced in the gland and thus given off in the milk are probably the same as those found in the blood if the animal were infected by conventional routes. Purification of the lactoglobulin protein fraction containing the antibody eliminated the non-specific inhibitors. These results were obtained from the various myxoviruses and mumps that propagated in the goat mammary gland.The main advantages of using the mammary gland as compared to using laboratory animals and their blood are as follows.1. Larger volumes of antibodies can be produced at one time (lactation period) against the influenza and mumps viruses for diagnostic and possibly for therapeutic uses.2. The animal (goat) does not appear to be affected whatsoever by the virus instillation and propagation techniques.3. The milk technique is therefore more humane towards laboratory animals. Invariably the laboratory animal does not have to be sacrificed.

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