Antigenic analysis of Pasteurella multocida (serotype 1) by crossed immunoelectrophoresis: characterization of whole cell associated antigens

1980 ◽  
Vol 26 (12) ◽  
pp. 1392-1402
Author(s):  
J. L. Bhasin ◽  
L. Lapointe-Shaw
Keyword(s):  
Pasteurella Multocida ◽  
Cell Envelope ◽  
Potassium Thiocyanate ◽  
Antigenic Analysis ◽  
Whole Cell ◽  
Heat Stable ◽  
Crossed Immunoelectrophoresis ◽  
Serotype 1 ◽  
Reference Preparation ◽  
Antigen Antibody

Crossed immunoelectrophoresis and other related quantitative immunoelectrophoretic techniques have been used to elucidate the antigenic complexity of a reference preparation of capsular extract, potassium thiocyanate extract, lipopolysaccharide, heat-stable antigens, and free endotoxin from Pasteurella multocida serotype 1. The reactions of these cellular fractions, in crossed immunoelectrophoresis, with reference anti-whole cell immunoglobulins disclosed five antigens in the capsular extract, seven in the potassium thiocyanate extract, one to three in the lipopolysaccharide, three in the heat-stable antigens, and five in the free endoxin. Comparison of these reference antigen–antibody systems, in crossed immunoelectrophoresis, with intermediate gel containing either a reference anti-cell envelope or anticytoplasmic immunoglobulins not only revealed the presence of additional antigens but also gave insight into the probable cellular origins (i.e., cell surface, cell envelope, or cytoplasm) of various antigens unveiled by reference anti-whole cell immunoglobulins. Using the principle of tandem crossed immunoelectrophoresis and crossed-line immunoelectrophoresis the immunochemical relationships between the antigenic components of these reference antigen–antibody systems were established.

Antigenic analysis of Pasteurella multocida (serotype 1) by crossed immunoelectrophoresis: characterization of cytoplasmic and cell envelope associated antigens

1980 ◽  
Vol 26 (6) ◽  
pp. 676-689
Author(s):  
J. L. Bhasin ◽  
L. Lapointe-Shaw
Keyword(s):  
Pasteurella Multocida ◽  
Cell Envelope ◽  
Rabbit Serum ◽  
Antigenic Analysis ◽  
Whole Cells ◽  
Crossed Immunoelectrophoresis ◽  
Serotype 1 ◽  
Determining Factor ◽  
Envelope Antigen

The application of crossed immunoelectrophoresis to the analysis of a reference cytoplasm and cell envelope preparation from Pasteurella multocida serotype 1 revealed antigenic complexity not previously found. At least 55 cytoplasmic and 19 cell envelope antigens were clearly distinguished. Variation of anticytoplasm immunoglobulin concentration was a major determining factor in resolving the maximum array of cytoplasmic antigens.The use of intermediate gel modification of crossed immunoelectrophoresis permitted the recognition of antibodies in the preimmune rabbit serum against a number of cytoplasmic antigens and a single cell envelope antigen. This technique also demonstrated that reference cytoplasm obtained by 105 000 × g centrifugation of sonically disrupted pasteurellae and repeatedly washed reference cell envelope preparation contained antigens of either origin in amounts sufficient to elicit antibody responses in the host. Antisera to whole cells in the intermediate gel indicated that formalin killed P. multocida were capable of eliciting immune responses to both reference systems.

Immunogenicity of potassium thiocyanate extracted and electrofocused Pasteurella multocida X-73I antigens in chickens and mice

1982 ◽  
Vol 28 (11) ◽  
pp. 1219-1225 ◽  
Author(s):  
Kevin L. McKinney ◽  
Paul A. Rebers ◽  
Richard B. Rimler
Keyword(s):  
Antibody Response ◽  
Pasteurella Multocida ◽  
Potassium Thiocyanate ◽  
Particulate Fraction ◽  
Supernatant Fraction ◽  
Chicken Embryos ◽  
Isoelectric Points ◽  
Serotype 1 ◽  
Soluble Supernatant

The immunogenicity of antigenic fractions obtained by the extraction of Pasteurella multocida strain X-73 (serotype 1) with potassium thiocyanate (KSCN) was determined in chickens and mice. The initial KSCN extract was centrifuged at 105 000 × g, and the antigens were separated into a particulate fraction (40p) and a soluble supernatant fraction (40s). The ultracentrifuged fractions were further resolved by preparative electrofocusing. The 40p fraction was resolved into two subgroups having isoelectric points of 3.5–3.9 and 5.5–6.0; the 40s fraction was resolved into five subgroups ranging in isoelectric points from 4.4 to 9.0. The 40p fractions were antigenically similar and contained lipopolysaccharide (LPS) and protein. The 40s fractions were antigenically distinct from the 40p fractions and from each other; they contained proteins and polysaccharides but no LPS. The 40p antigens were strongly immunogenic in mice and chickens, whereas the 40s antigens were weakly immunogenic in chickens and not immunogenic in mice. The incorporation of Freund's complete adjuvant increased the immunogenicity of the 40s antigens in chickens. The 40p antigens induced greater frequencies of serological responses in chickens than the 40s antigens as detected by counterimmunoelectrophoresis and immunodiffusion. This suggested that the increased protection associated with the 40p antigens may have been the result of better antibody response. The toxicity of all the fractions was evaluated by determination of lethality for 10-day-old chicken embryos because of the sensitivity and reliability of the test. The 40p fraction had an LD50 = 0.38 μg, and the 40s fraction had an LD50 = 2.5 μg. Since the 40s fraction contained no detectable LPS, it is likely that two toxins are present, one which contains LPS and one which does not.

Permeability Through Bacterial Porins Dictates Whole Cell Compound Accumulation

2020 ◽  
Author(s):  
Silvia Acosta Gutiérrez ◽  
Igor Bodrenko ◽  
Matteo Ceccarelli
Keyword(s):  
Cell Envelope ◽  
Scoring Function ◽  
Modern Medicine ◽  
New Drugs ◽  
Prediction Ability ◽  
Whole Cell ◽  
Virtual Libraries ◽  
Major Bottleneck ◽  
Global Threat ◽  
Bacterial Porins

The lack of new drugs for Gram-negative pathogens is a global threat to modern medicine. The complexity of their cell envelope, with an additional outer membrane, hinders internal accumulation and thus, the access of molecules to targets. Our limited understanding of the molecular basis for compound influx and efflux from these pathogens is a major bottleneck for the discovery of effective antibacterial compounds. Here we analyse the correlation between the whole-cell compound accumulation of ~200 molecules and their predicted porin permeability coefficient (influx), using a recently developed scoring function. We found a strong linear relationship (75%) between the two, confirming porins key role in compound penetration. Further, the remarkable prediction ability of the scoring function demonstrates its potentiality to guide the optimization of hits to leads as well as the possibility of screening ultra-large virtual libraries. Eventually, the analysis of false positives, molecules with high-predicted influx but low accumulation, provides new hints on the molecular properties behind efflux.<br>

THE EFFECT OF BACTERIAL CELL LYSIS AND OF PLANT EXTRACTS ON CELLULOSE PRODUCTION BY ACETOBACTER XYLINUM

1963 ◽  
Vol 41 (1) ◽  
pp. 1691-1702 ◽  
Author(s):  
T. E. Webb ◽  
J. Ross Colvin
Keyword(s):  
Bacterial Cell ◽  
Cell Envelope ◽  
Acetobacter Xylinum ◽  
Cellulose Synthesis ◽  
Additive Effects ◽  
Cellulose Production ◽  
Whole Cells ◽  
Negative Results ◽  
Heat Stable ◽  
Plant Sources

The production of cellulose by lysozyme lysates of Acetobacter xylinum is similar to that of a suspension of whole cells, in contrast to the negative results obtained with previous "cell-free" preparations. The results of differential centrifugation of these lysates suggests that most of the enzymes required for cellulose synthesis from glucose normally are held by the cell envelope and are not located in the cytoplasm. However, a heat-stable cofactor(s) is present in the supernatant derived from the cell contents which may stimulate cellulose synthesis by the cell envelopes.The addition of extracts from a number of plant sources increased cellulose synthesis by whole cells of A. xylinum. In particular, the supernatant prepared by centrifugation of an homogenate of tomatoes increased bacterial cellulose production at pH 6 by a factor of 3. Both dialyzable and non-dialyzable substances in the extract are responsible. Fractionation of the non-dialyzable portion of the extract by column chromatography suggests that the overall increase is due to additive effects of several compounds. Here also the compounds appear to act upon the bacterial cell envelope.

Rate nephelometric measurement of rheumatoid factor in serum.

1979 ◽  
Vol 25 (11) ◽  
pp. 1909-1914 ◽  
Author(s):  
P R Finley ◽  
M J Hicks ◽  
R J Williams ◽  
J Hinlicky ◽  
D A Lichti
Keyword(s):  
Rheumatoid Factor ◽  
Control Level ◽  
Positive Control ◽  
Standard Curve ◽  
Graph Paper ◽  
Rate Of Increase ◽  
Human Sera ◽  
Reference Preparation ◽  
Level I ◽  
Antigen Antibody

Abstract We describe the measurement of rheumatoid factor in human sera with a rate nephelometer. The National Reference Preparation for Rheumatoid Factors is used to calibrate the assay in International Units. We used Hyland Positive Control, Level I, as a secondary standard. The standard curve is exponential, but is linear when plotted on log-log graph paper. Aggregated immune globulin (IgG) is the antigen used to detect rheumatoid factor (IgM-class antibody to IgG). The rate reaction measures the rate of increase in light-scatter by the antigen-antibody complexes; the reaction takes place in 17 to 20 s. Precision, linearity, and accuracy are excellent. Results agree well with those for a commonly used latex precipitation test. The advantages of speed, quantification in International Units, and superior discrimination of concentration as compared to serological titration provide a more reliable test for use in the diagnosis and treatment of rheumatoid arthritis.

scholarly journals BIOCHEMICAL STUDY OF CELLULAR ANTIGEN-ANTIBODY REACTION IN TISSUE CULTURE

1960 ◽  
Vol 112 (2) ◽  
pp. 249-255 ◽  
Author(s):  
Akira Tokuda ◽  
Hideo Hayashi ◽  
Kinishiro Matsuba
Keyword(s):  
Tissue Culture ◽  
Protease Activity ◽  
Biochemical Study ◽  
Culture Fluid ◽  
Partial Purification ◽  
Trichloracetic Acid ◽  
Heat Stable ◽  
Antibody Reaction ◽  
Antigen Antibody ◽  
Cellular Antigen

Decrease in the protease activity of the culture fluid observed at later stages of the antigen-antibody reaction is believed to be due to the release of an inhibitor by the cells. The inhibitor was submitted to partial purification: it is heat-stable, non-precipitated by trichloracetic acid and non-dialyzable. It inhibits certain cellular and tissue proteases and papain but is inactive against trypsin. It is suggested that the balance between protease and anti-protease released may determine the intensity, extent, and duration of certain sensitization phenomena.

Enzymes and Cells Confined in Silica Nanopores

2000 ◽  
Vol 651 ◽  
Author(s):  
Jacques Livage ◽  
Cécile Roux ◽  
Thibaud Coradin ◽  
Souad Fennouh ◽  
Stéphanie Guyon ◽  
...  
Keyword(s):  
Sol Gel ◽  
Silica Gels ◽  
Cellular Organization ◽  
Silica Network ◽  
Whole Cell ◽  
Molecular Precursors ◽  
Measured Activity ◽  
Human Sera ◽  
Gel Glasses ◽  
Antigen Antibody

AbstractThe sol-gel process opens new possibilities in the field of biotechnologies. Sol-gel glasses are formed at room temperature via the polymerization of molecular precursors. Enzymes can be added to the solution of precursors and trapped within the growing silica network. Small substrate molecules can diffuse through the pores allowing reactions to be performed in-situ, within the silica gels. Enzyme are encased by the hydrated silica in a cage tailored to their size, they retain their biocatalytic activity and may even be stabilized within the sol-gel matrix.Whole cell bacteria have also been immobilized within sol-gel glasses. They behave as a "bag of enzymes" and their membrane protects enzymes against denaturation and leaching. The cellular organization of bacteria cells is preserved upon encapsulation. Experiments performed with Escherichia coli induced to β-galactosidase show that they still exhibit noticeable enzymatic activity. Some degradation of the cell walls may even occur increasing the “measured” activity. However silica gels made from aqueous precursors seem to prevent bacteria from natural degradation upon ageing.Antibody-antigen recognition has been shown to be feasible within sol-gel matrices. Trapped antibodies bind specifically the corresponding haptens and can be used for the detection of traces of chemicals. Even whole cell protozoa have been encapsulated without any alteration of their cellular organization. For medical applications, trapped parasitic protozoa have been used as antigens for blood tests with human sera. Antigen-antibody interactions were followed by the so-called Enzyme Linked ImmunoSorbent Assays (ELISA).

scholarly journals Studies on the Nature of the Circulating Anticoagulant Directed against Antihemophilic Factor: With Notes on an Assay for Antihemophilic Factor

Blood ◽  
1962 ◽  
Vol 20 (2) ◽  
pp. 137-149 ◽  
Author(s):  
ROBERT T. BRECKENRIDGE ◽  
OSCAR D. RATNOFF
Keyword(s):  
Substrate Concentration ◽  
Clotting Time ◽  
Heat Stable ◽  
Concentration Dependency ◽  
Proportional Relationship ◽  
Kaolin Clotting Time ◽  
Antigen Antibody ◽  
Kinetics Of ◽  
Temperature Time ◽  
Antihemophilic Factor

Abstract Modification of the kaolin clotting time has produced a reliable, simple assay system for antihemophilic factor. This system has been utilized for an investigation of the nature and mode of action of the circulating anticoagulant directed against antihemophilic factor. The anticoagulant has been shown to be present in equal amounts in plasma and serum, to be associated with fractions II and III of plasma prepared by the method of Cohn, to be relatively heat stable and to be stable for prolonged periods at -20 C. Investigation of the kinetics of the anticoagulant-antihemophilic reaction has demonstrated its temperature, time, pH, and substrate concentration dependency. The anticoagulant is not inactivated during the reaction with antihemophilic factor, and there is a proportional relationship between the anticoagulant and the antihemophilic factor during their interaction. No antigen-antibody manifestations could be detected during the anticoagulant-antihemophilic factor reaction. These characteristics support the hypothesis that the inactivation of antihemophilic factor by specific circulating anticoagulants is enzymatic.

scholarly journals Generation and Molecular Characterization of New Temperature-Sensitive Plasmids Intended for Genetic Engineering of Pasteurellaceae

2005 ◽  
Vol 71 (11) ◽  
pp. 7187-7195 ◽  
Author(s):  
Robert E. Briggs ◽  
Fred M. Tatum
Keyword(s):  
Base Pair ◽  
Pasteurella Multocida ◽  
Chemical Mutagenesis ◽  
Temperature Sensitive ◽  
Origin Of Replication ◽  
Wild Type ◽  
Recognition Sequence ◽  
Nucleotide Substitutions ◽  
Serotype 1

ABSTRACT Temperature-sensitive (TS) plasmids were generated through chemical mutagenesis of a derivative of the streptomycin resistance parent plasmid pD70, isolated from Mannheimia hemolytica serotype 1. Three TS plasmids which failed to replicate at or above 42°C in M. hemolytica but which were fully functional below 31°C were selected for further analysis. Two of the TS plasmids were shown by sequencing to possess unique single-base-pair mutations. The third TS plasmid contained a unique base pair substitution and a second mutation that had been previously identified. These mutations were clustered within a 200-bp region of the presumed plasmid origin of replication. Site-directed single-nucleotide substitutions were introduced into the wild-type pD70 origin of replication to confirm that mutations identified by sequencing had conferred thermoregulated replication. Deletion analysis on the wild-type pD70 plasmid replicon revealed that approximately 720 bp are necessary for plasmid maintenance. Replication of the TS plasmids was thermoregulated in Pasteurella multocida and Haemophilus somnus as well. To consistently transform H. somnus with TS plasmid, in vitro DNA methylation with commercially available HhaI methyltransferase was necessary to protect against the organism's restriction enzyme HsoI (recognition sequence 5′-GCGC-3′) characterized herein.

Sign in / Sign up

Export Citation Format

Share Document