scholarly journals AUTORADIOGRAPHIC DETECTION OF ANTIGENS IN CELLS USING TRITIUM-LABELED ANTIBODIES

1970 ◽  
Vol 18 (7) ◽  
pp. 490-497 ◽  
Author(s):  
K. OSTROWSKI ◽  
E. A. BARNARD ◽  
W. SAWICKI ◽  
T. CHORZELSKI ◽  
A. LANGNER ◽  
...  
Keyword(s):  
Acetic Anhydride ◽  
Fluorescent Marker ◽  
Isotopic Method ◽  
Relative Quantitation ◽  
Antibody Titers ◽  
Autoradiographic Detection ◽  
Very High ◽  
In Cells

Immunoglobulins specific for cellular antigens of several types were treated with 3H-acetic anhydride in conditions which were shown to introduce three to four 3H-acetyl groups per molecule, with very high retention of the antibody titers. Methods were established for application of these labeled globulins in an autoradiographic procedure. The results show that localization of cell-bound antigens can be demonstrated by this method, by silver grain concentrations at sites that correspond to those demonstrated by the same globulins labeled with a fluorescent marker. A control nonimmune globulin was reacted similarly and gave no labeling of the same tissues. The advantage of the isotopic method would be the relative quantitation that it is, in principle, capable of yielding.

scholarly journals Assessment of 3D MINFLUX data for quantitative structural biology in cells

2021 ◽  
Author(s):  
Kirti Prakash ◽  
Alistair Curd
Keyword(s):  
Quantitative Analysis ◽  
Structural Biology ◽  
Single Molecule ◽  
Structural Features ◽  
Localization Microscopy ◽  
Current State ◽  
New Development ◽  
Very High ◽  
Single Molecule Localization Microscopy ◽  
In Cells

MINFLUX is a promising new development in single-molecule localization microscopy, claiming a resolution of 1-3 nm in living and fixed biological specimens. While MINFLUX can achieve very high localisation precision, quantitative analysis of reported results leads us to dispute the resolution claim and question reliability for imaging sub-100-nm structural features, in its current state.

scholarly journals Global Absence and Targeting of Protective Immune States in Severe COVID-19.

2020 ◽  
Author(s):  
Matthew Krummel ◽  
Alexis Combes ◽  
Tristan Courau ◽  
Nicholas Kuhn ◽  
kenneth Hu ◽  
...  
Keyword(s):  
Single Cell Analysis ◽  
Severe Disease ◽  
Cell Types ◽  
Cell Analysis ◽  
Lower Viral Load ◽  
Mild Disease ◽  
Holistic Understanding ◽  
Antibody Titers ◽  
Very High ◽  
Analysis Protocol

Abstract While SARS-CoV-2 infection has pleiotropic and systemic effects in some patients, many others experience milder symptoms. We sought a holistic understanding of the severe/mild distinction in COVID-19 pathology, and its origins. We performed a wholeblood preserving single-cell analysis protocol to integrate contributions from all major cell types including neutrophils, monocytes, platelets, lymphocytes and the contents of serum. Patients with mild COVID-19 disease display a coordinated pattern of interferonstimulated gene (ISG) expression across every cell population and these cells are systemically absent in patients with severe disease. Severe COVID-19 patients also paradoxically produce very high anti-SARS-CoV-2 antibody titers and have lower viral load as compared to mild disease. Examination of the serum from severe patients demonstrates that they uniquely produce antibodies with multiple patterns of specificity against interferon-stimulated cells and that those antibodies functionally block the production of the mild disease-associated ISG-expressing cells. Overzealous and autodirected antibody responses pit the immune system against itself in many COVID-19 patients and this defines targets for immunotherapies to allow immune systems to provide viral defense.

scholarly journals THE PROVOCATION OF LATENT LYMPHOCYTIC CHORIOMENINGITIS VIRUS INFECTIONS IN MICE BY TREATMENT WITH ANTILYMPHOCYTIC SERUM

1968 ◽  
Vol 127 (2) ◽  
pp. 327-339 ◽  
Author(s):  
Mogens Volkert ◽  
Claus Lundstedt
Keyword(s):  
Considerable Increase ◽  
Latent Infection ◽  
Lymphocytic Choriomeningitis ◽  
Antilymphocytic Serum ◽  
Virus Infections ◽  
Adult Mice ◽  
Antibody Titers ◽  
Sublethal Doses ◽  
High Virus ◽  
Very High

The hypothesis that treatment with antilymphocytic serum (ALS) can provoke latent virus infections has been investigated. In adult mice infections with sublethal doses of LCM virus usually result in the development of immunity to the virus and at the same time to a prolonged latent infection. In the experiments described an intensive treatment with large doses of ALS was given to mice which had recovered from LCM virus infection. At the beginning of the treatment the mice had high titers of complement-fixing antibodies in their blood and no detectable virus. The data presented show that in spite of the immunity the ALS treatment provoked the occult virus and led to the development of viremia in all the treated mice. In some, very high virus titers were demonstrable. When the ALS treatment was discontinued the viremia disappeard again. In most of the mice the ALS did not suppress the complement-fixing antibody titers and in some there was even a considerable increase in titer. In such cases the increases in virus titers and in antibody titers were closely related to one another. These results demonstrate once again that the complement-fixing antibodies to the LCM virus in mice probably do not influence the virus.

scholarly journals Differences of antigenic profiles on immunoblotting of wild type measles virus and vaccine virus in Indonesia

2016 ◽  
Vol 48 (6) ◽  
pp. 364
Author(s):  
Made Setiawan ◽  
Agus Sjahrurachman ◽  
Fera Lbrahim ◽  
Agus Suwandono
Keyword(s):  
Measles Virus ◽  
Vaccine Virus ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Mmr Vaccine ◽  
N Protein ◽  
Antibody Titers ◽  
Antigen Antibody ◽  
Very High

Background Measles virus has a negative, single strand RNAgenome which codes for six important structural proteins. Thegenes of the wild type measles virus have many variances hencethe nucleotide sequences of each wild type virus and vaccine virusare different. This differences lead to the antigenic differencesbetween wild type and vaccine virus.Objective The purpose of this research is to investigate thedifferences in the antigenic profiles on immunoblotting betweenwild type and vaccine virus.Results The analysis results are 1) the antigen ofCAM-70 vaccinevirus was less able in cross reacting with the antibodies from G2,G3, 09, CAM-70 and Schwarz; 2) The antibody aga inst CAM-70 was only able to cross react with antigens of N protein and afew of antigens ofF proteins; 3) The wild type virus were veryimmunogenic, hence the antibody titers were very high; 4) TheCAM-70 and MMR vaccine virus were less immunogenic, hencetheir antibody were very low; 5) The antibody responses thatalways occurred from all immunized mice serum were antibodyfor N and F proteins. However, the antibody against CAM-70vaccine virus was still able to react with wild type virus (G2, G3and 09).Conclusion All antigen-antibody reaction on immunoblottingresulted in different profiles especially between wild type virusand CAM-70 vaccine virus. Although CAM-70 vaccine virusshowed clear differences compared to G2, G3 and 09 genotypes,antibodies against CAM-70 were still able to cross react withantigens from other genotypes (G2, G3 and D9).

scholarly journals Potent neutralizing equine antibodies raised against recombinant SARS-CoV-2 spike protein for COVID-19 passive immunization therapy

2020 ◽  
Author(s):  
Luis Eduardo R. Cunha ◽  
Adilson A. Stolet ◽  
Marcelo A. Strauch ◽  
Victor A. R. Pereira ◽  
Carlos H. Dumard ◽  
...  
Keyword(s):  
Clinical Trial ◽  
Therapeutic Intervention ◽  
Industrial Production ◽  
Serum Antibody ◽  
Passive Immunization ◽  
Neutralizing Antibody ◽  
Spike Protein ◽  
Safety And Efficacy ◽  
Antibody Titers ◽  
Very High

AbstractWe used the trimeric spike (S) glycoprotein (residues 1-1208) in the prefusion conformation to immunize horses for production of hyperimmune globulins against SARS-CoV-2. Serum antibody titers measured by anti-spike ELISA were above 1:1,000,000, and neutralizing antibody titer was 1:14,604 (average PRNT90), which is 140-fold higher than the average neutralizing titer of plasma from three convalescent COVID-19 patients analyzed for comparison. Using the same technology routinely used for industrial production of other horse hyperimmune products, plasma from immunized animals was pepsin digested to remove the Fc portion and purified, yielding a F(ab’)2 preparation with PRNT90 titers 150-fold higher than the neutralizing titers in human convalescent plasma. Repeating the hyperimmunization in a second group of horses confirmed the very high neutralizing titers in serum and in a GMP clinical F(ab’)2 lot. Virus-neutralizing activity in samples from mice that received the F(ab’)2 preparation was detected even three days after injection, indicating an appropriate half-life for therapeutic intervention. These results supported the design of a clinical trial (identifier NCT04573855) to evaluate safety and efficacy of this horse F(ab’)2 preparation.

scholarly journals Studies on the, Comparative Physiology of Chara Australis 1. Growth Pattern and Gross Cytology of the Internodal Oell

1964 ◽  
Vol 17 (1) ◽  
pp. 49 ◽  
Author(s):  
Marie J Peebles FV Mercer ◽  
TC Chambers
Keyword(s):  
Growth Pattern ◽  
Unit Volume ◽  
Exponential Phase ◽  
Linear Phase ◽  
Comparative Physiology ◽  
Growth Phases ◽  
Internodal Cell ◽  
Chara Australis ◽  
Very High ◽  
In Cells

The growth of the internodal cell and its parts appears to be allometric. Two growth phases are recognized, an exponential phase in the young cells followed by a linear phase during which the internode expands by up to 200 mm in length. In the exponential phase the cytoplasm to vacuole ratio declines from a very high value to about 0�04--0�06 and then remains constant throughout the linear phase. In the linear phase the structure of the protoplast, at the light.microscope level, is similar for all cells; a unit volume of the protoplast appears similar in cells from 10 to 200 rum in length.

scholarly journals Global Absence and Targeting of Protective Immune States in Severe COVID-19

2020 ◽  
Author(s):  
Alexis J. Combes ◽  
Tristan Courau ◽  
Nicholas F. Kuhn ◽  
Kenneth H. Hu ◽  
Arja Ray ◽  
...  
Keyword(s):  
Immune System ◽  
Single Cell Analysis ◽  
Severe Disease ◽  
Cell Types ◽  
Mild Disease ◽  
Holistic Understanding ◽  
Antibody Titers ◽  
Very High ◽  
Analysis Protocol ◽  
Successful Production

AbstractWhile SARS-CoV-2 infection has pleiotropic and systemic effects in some patients, many others experience milder symptoms. We sought a holistic understanding of the severe/mild distinction in COVID-19 pathology, and its origins. We performed a whole-blood preserving single-cell analysis protocol to integrate contributions from all major cell types including neutrophils, monocytes, platelets, lymphocytes and the contents of serum. Patients with mild COVID-19 disease display a coordinated pattern of interferon-stimulated gene (ISG) expression across every cell population and these cells are systemically absent in patients with severe disease. Severe COVID-19 patients also paradoxically produce very high anti-SARS-CoV-2 antibody titers and have lower viral load as compared to mild disease. Examination of the serum from severe patients demonstrates that they uniquely produce antibodies with multiple patterns of specificity against interferon-stimulated cells and that those antibodies functionally block the production of the mild disease-associated ISG-expressing cells. Overzealous and auto-directed antibody responses pit the immune system against itself in many COVID-19 patients and this defines targets for immunotherapies to allow immune systems to provide viral defense.One Sentence SummaryIn severe COVID-19 patients, the immune system fails to generate cells that define mild disease; antibodies in their serum actively prevents the successful production of those cells.

scholarly journals Epitope Mapping Porcine Reproductive and Respiratory Syndrome Virus by Phage Display: the nsp2 Fragment of the Replicase Polyprotein Contains a Cluster of B-Cell Epitopes

2001 ◽  
Vol 75 (7) ◽  
pp. 3277-3290 ◽  
Author(s):  
M. B. Oleksiewicz ◽  
A. Bøtner ◽  
P. Toft ◽  
P. Normann ◽  
T. Storgaard
Keyword(s):  
Phage Display ◽  
B Cell ◽  
Immunoglobulin A ◽  
Respiratory Syndrome Virus ◽  
B Cell Epitopes ◽  
Protein Fragments ◽  
Phage Display Libraries ◽  
Antibody Titers ◽  
Very High

ABSTRACT We screened phage display libraries of porcine reproductive and respiratory syndrome virus (PRRSV) protein fragments with sera from experimentally infected pigs to identify linear B-cell epitopes that are commonly recognized during infection in vivo. We identified 10 linear epitope sites (ES) 11 to 53 amino acids in length. In the replicase polyprotein, a total of eight ES were identified, six of which localized to the Nsp2 replicase polyprotein processing end product. In the structural proteins, a total of two ES were identified, in the ORF3 and ORF4 minor envelope glycoproteins. The ORF4 ES was previously identified by monoclonal antibody mapping (J. J. M. Meulenberg, A. P. van Nieuwstadt, A. van Essen-Zandenbergen, and J. P. M. Langeveld, J. Virol. 71:6061–6067, 1997), but its immunogenicity had not been examined in pigs. We found that six experimentally PRRSV-infected pigs consistently had very high antibody titers against the ORF4 ES. In some animals, sera diluted 1:62,500 still gave weak positive enzyme immunoassay reactivity against the ORF4 ES. This hitherto unrecognized immunodominance likely caused phages displaying the ORF4 ES to outcompete phages displaying other ES during library screening with porcine sera and accounted for our failure to identify more than two ES in the structural genes of PRRSV. Genetic analysis showed that variable ES were also the most immunogenic in vivo. Serological analysis indicated differences in the immunoglobulin A responses between short-term and longer-term viremic pigs towards some ES. The implications of these findings for PRRSV diagnostics and immunopathogenesis are discussed.

scholarly journals Dihydrorhodamine 123 identifies impaired mitochondrial respiratory chain function in cultured cells harboring mitochondrial DNA mutations.

1996 ◽  
Vol 44 (6) ◽  
pp. 571-579 ◽  
Author(s):  
C Sobreira ◽  
M Davidson ◽  
M P King ◽  
A F Miranda
Keyword(s):  
Mitochondrial Dna ◽  
Respiratory Chain ◽  
Mitochondrial Function ◽  
Cultured Cells ◽  
Mitochondrial Respiratory Chain ◽  
Mtdna Mutations ◽  
Dihydrorhodamine 123 ◽  
Very High ◽  
Mitochondrial Respiratory ◽  
In Cells

Several human diseases have been found to be caused by mitochondrial DNA (mtDNA) mutations. Pathogenic mutated (mut) mtDNAs are usually "heteroplasmic," coexisting intracellularly with wild-type (wt) mtDNAs. For some mtDNA mutations, cells have normal levels of respiratory chain function unless the percentage of mut-mtDNA is very high. Although progress in understanding the molecular basis of mitochondrial diseases has been remarkable, the heterogeneity of mut-mtDNA distribution, even among cells of the same tissue, makes it difficult to clearly delineate the relationships between mtDNA mutations, gene dosage, and clinical phenotypes. In a search for screening methods for identifying cultured cells with deficient mitochondrial function, we incubated living cells harboring mut-mtDNAs with dihydrorhodamine 123 (DHR123), an uncharged, nonfluorescent agent that can be converted by oxidation to the fluorescent laser dye rhodamine 123 (R123). Bright mitochondrial staining was observed in cells that respired normally. Fluorescence was significantly reduced in cells with mitochondrial respiratory chain dysfunction resulting from very high levels of mut-mtDNAs. The data show that DHR123 is useful for assessing mitochondrial function in single cells, and can be used for isolating viable, respiratory chain-deficient cells from heterogeneous cultures.

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