scholarly journals Luminescent detection method for immunodot, Western, and Southern blots.

1986 ◽  
Vol 34 (12) ◽  
pp. 1645-1650 ◽  
Author(s):  
M M Leong ◽  
C Milstein ◽  
R Pannell
Keyword(s):  
Monoclonal Antibody ◽  
Cell Line ◽  
Indirect Method ◽  
Detection Method ◽  
Specific Activity ◽  
High Specific Activity ◽  
The Other ◽  
X Ray ◽  
Bispecific Monoclonal Antibody ◽  
Direct Binding

An anti-peroxidase-anti-biotin hybrid hybridoma rat cell line, capable of producing a bispecific monoclonal antibody, has been derived to explore its use in conjunction with a luminol immunodetection system. Luminescence was detected using x-ray film. The method was sufficiently sensitive and effective, but was less sensitive than autoradiographic methods using high-specific-activity 32P-labeled probes. Exposure times, on the other hand, were of the order of seconds rather than days. The direct binding of both peroxidase and biotin by the bispecific monoclonal antibody is simpler but less sensitive than the more conventional indirect method using a commercial peroxidase coupled with anti-rat antibody as a developing antibody.

Hybridoma Screening by Antigen Capture: Capture or Sandwich ELISA in 96-Well Plates

2022 ◽  
Vol 2022 (1) ◽  
pp. pdb.prot103127
Author(s):  
Edward A. Greenfield
Keyword(s):  
Culture Supernatant ◽  
Detection Method ◽  
Specific Activity ◽  
Binding Capacity ◽  
Sandwich Elisa ◽  
High Specific Activity ◽  
Tissue Culture Supernatant ◽  
Capture Assay ◽  
Antigen Capture ◽  
Elisa Plate

In an antigen capture assay for hybridoma screening, the detection method identifies the presence of the antigen. Often this is achieved by labeling the antigen directly. In this assay, the polyvinyl chloride (PVC) wells of a high-binding-capacity ELISA plate are first coated with an affinity-purified rabbit anti-mouse immunoglobulin and then incubated with hybridoma tissue culture supernatant. Monoclonal antibodies in the supernatant are “captured” on the coated PVC surface and detected by screening with biotin- or histidine (His)–tagged antigen. The antigen can be labeled to a high specific activity and thus very little antigen is required for this procedure.

scholarly journals Preliminary X-ray diffraction analysis of a thermophilic β-1,3–1,4-glucanase fromClostridium thermocellum

2014 ◽  
Vol 70 (7) ◽  
pp. 946-948 ◽  
Author(s):  
Lilan Zhang ◽  
Puya Zhao ◽  
Chun-Chi Chen ◽  
Chun-Hsiang Huang ◽  
Tzu-Ping Ko ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Specific Activity ◽  
High Specific Activity ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Glycosidic Bonds ◽  
Hydrolysis Of

β-1,3–1,4-Glucanases catalyze the specific hydrolysis of internal β-1,4-glycosidic bonds adjacent to the 3-O-substituted glucose residues in mixed-linked β-glucans. The thermophilic glycoside hydrolase CtGlu16A fromClostridium thermocellumexhibits superior thermal profiles, high specific activity and broad pH adaptability. Here, the catalytic domain of CtGlu16A was expressed inEscherichia coli, purified and crystallized in the trigonal space groupP3121, with unit-cell parametersa=b= 74.5,c= 182.9 Å, by the sitting-drop vapour-diffusion method and diffracted to 1.95 Å resolution. The crystal contains two protein molecules in an asymmetric unit. Further structural determination and refinement are in progress.

scholarly journals X-Ray Powder Diffraction of Einsteinium Compounds

1978 ◽  
Vol 22 ◽  
pp. 101-107
Author(s):  
R. G. Haire ◽  
J. R. Peterson
Keyword(s):  
Chemical Composition ◽  
Powder Diffraction ◽  
Thermal Energy ◽  
Diffraction Data ◽  
Specific Activity ◽  
Radioactive Decay ◽  
High Specific Activity ◽  
X Ray ◽  
Transplutonium Elements ◽  
Xrpd Analysis

X-ray powder diffraction (XRPD) analysis has proven to be extremely valuable in the study of the transplutonium elements. The high specific activity and the thermal energy associated with the radioactive decay of einsteinium isotopes make XRPD analysis of einsteinium samples very difficult. The major problems are destruction of the samples crystallinity and alteration of their chemical composition. Blackening of the X-ray film also compromises the analyses. By applying certain guidelines given here, a limited amount of success has been achieved in obtaining diffraction data on einsteinium and some of its compounds.

Monoclonal antibody-dendrimer conjugates enable radiolabeling of antibody with markedly high specific activity with minimal loss of immunoreactivity

2000 ◽  
Vol 27 (9) ◽  
pp. 1334-1339 ◽  
Author(s):  
Hisataka Kobayashi ◽  
Noriko Sato ◽  
Tsuneo Saga ◽  
Yuji Nakamoto ◽  
Takayoshi Ishimori ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Specific Activity ◽  
High Specific Activity ◽  
Minimal Loss ◽  
Dendrimer Conjugates

SKELETAL SILVER CATALYSTS FOR THE OXIDATION OF ETHYLENE TO ETHYLENE OXIDE

1956 ◽  
Vol 34 (5) ◽  
pp. 665-671 ◽  
Author(s):  
A. Cambron ◽  
W. A. Alexander
Keyword(s):  
Ethylene Oxide ◽  
Alkaline Earth ◽  
Specific Activity ◽  
High Specific Activity ◽  
Alkaline Earth Metals ◽  
The Other ◽  
Silver Alloys ◽  
Silver Catalysts

Skeletal silver catalysts of high specific activity have been prepared by the removal of calcium from calcium–silver alloys. The activity of these catalysts in the oxidation of ethylene to ethylene oxide has been investigated. Catalysts prepared by the removal of the other alkaline earth metals from their alloys with silver have also been studied. It has been found that catalysts prepared from calcium–silver alloys show a higher specific activity and are more stable and more conveniently prepared than the catalysts from the other silver alloys investigated.

The First Human Cell Line Recombinant Factor VIII (Human-cl rhFVIII) - Functional Properties and Differences From Currently Available Recombinant Factor VIII Products.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1298-1298
Author(s):  
Bruce A. Schwartz ◽  
Helena Sandberg ◽  
Irene Agerkvist ◽  
Christoph Kannicht ◽  
Margareta Ramström ◽  
...  
Keyword(s):  
Factor Viii ◽  
Cell Line ◽  
Functional Properties ◽  
Human Cell ◽  
Specific Activity ◽  
Activated Protein C ◽  
High Specific Activity ◽  
Human Cell Line ◽  
Recombinant Factor Viii ◽  
Chromogenic Assay

Abstract Abstract 1298 Poster Board I-320 All current recombinant factor VIII products produced today are derived from the expression of FVIII in hamster cell lines. This is a report of the functional properties of a newly developed recombinant factor VIII concentrate produced in a human cell line. The possible advantage of using a human cell line for expression of recombinant factor VIII is that a human pattern of post-translational modifications (PTM), such as glycosylation, are obtained. These human like PTMs, may possibly lead to improved function and a reduced immunogenic product profile compared to recombinant factor VIII products from CHO or BHK cells that have hamster-specific PTM. Data are presented on the characteristics of the Human-cl rhFVIII, a B-domain-deleted factor VIII protein produced in HEK293F cells, and how they compare with those of other factor VIII products. Human-cl rhFVIII is highly pure (>99.9% purity) with a high specific activity of 8000-11000 IU/mg protein, and a FVIII:C to FVIII:Ag ratio close to 1.0. This indicates that the entire factor VIII protein in the product is active. Human-cl rhFVIII showed no significant assay discrepancy between one-stage and chromogenic assay results. Because of the human glycosylation profile of the new Human-cl rhFVIII, there was an absence of the antigenic type of sialic acid, N-glycolylneuraminic acid as well as of antigenic Galα (1,3) Gal epitopes found in the non humane derived products. Another type of PTM of factor VIII is the sulfation of certain tyrosines that are crucial for the biological function e.g. the interaction with von Willebrand factor (vWF). Human-cl rhFVIII was found to be optimally sulfated when compared to endogenous human FVIII. In this context, it is of interest to note that Human cl-rhFVIII showed a 60% higher affinity in binding to vWF compared to other recombinant factor VIII products and the number of molecules in the product able to bind to vWF was significantly higher for Human-cl rhFVIII. Interaction with thrombin and capacity to generate thrombin were shown to be similar to those of plasma-derived factor VIII. The sensitivity to be inactivated by activated protein C was similar to all currently licensed rhFVIII products. The important properties of Human cell line derived recombinant factor VIII may be very promising for its use in future treatment of Haemophilia A. Disclosures Schwartz: Octapharma: Employment. Sandberg:Octapharma: Employment. Agerkvist:Octapharma: Employment. Kannicht:Octapharma: Employment. Ramström:Octapharma: Employment. Stenlund:Octapharma: Employment. Oswaldsson:Octapharma: Employment. Brunberg:Octapharma: Employment.

Fragment D Immunoreactivity: The Influence of Iodination and Calcium Environment

1979 ◽  
Author(s):  
B. Kudryfc ◽  
N. Blombäck
Keyword(s):  
Conformational Changes ◽  
Specific Antibody ◽  
Binding Sites ◽  
Specific Activity ◽  
High Specific Activity ◽  
The Other ◽  
Antigenic Determinants ◽  
Affinity Binding ◽  
Compact Structure ◽  
Chloramine T

Human fragment D (Fg-Ds) has been iodinated using both the Chloramine-T and lactoperoxidase methods. The specific activity was similar regardless of the method used. However, binding to a specific antibody was different for each preparation. The antigen labeled by the Chloramine-T method bound to a maximum of 40%, the other labeled product bound up to 85%. A correlation between the degree of immunoreactivity and avidity for a fibrinmonomer conjugate was found also. Fibrinmonomer bound about twice the amount of lactoperoxidase iodinated Fg-Ds as it did the Chloramine-T product. The use of these conjugates in the purification of immunoreactive Fg-Ds of high specific activity will be discussed. affinity binding sites for calcium have recently been demonstrated in fibrinogen. The presence of bound calcium is also believed to protect Fg-Ds from further digestion by plasmin. This is probably due to the formation of a more compact structure. However, conformational changes for calcium bound fibrinogen or Fg-Ds have not been observed. We tested the immunoreactivity of the lactoperoxidase iodinated Fg-Ds in presence and absence of calcium. Differences were found and this data suggests that some modification of antigenic determinants takes place as a consequence of calcium in the environment.

Behavior of radiozinc in rat plasma

1962 ◽  
Vol 202 (5) ◽  
pp. 926-930 ◽  
Author(s):  
J. P. Okunewick ◽  
B. Pond ◽  
T. G. Hennessy
Keyword(s):  
Intravenous Injection ◽  
Plasma Protein ◽  
Specific Activity ◽  
Zinc Ion ◽  
Rat Plasma ◽  
Direct Proportion ◽  
X Ray ◽  
Inorganic Ion ◽  
Direct Binding ◽  
Major Alteration

This series of studies was undertaken to determine the behavior of Zn65 when injected intravenously as the inorganic ion. A clearance curve for Zn65 similar to those of other workers was obtained. It was found that the decrease in clearance and distribution of Zn65 was not in direct proportion to the decrease in specific activity. Upon extraction of loosely bound zinc from plasma with dithizone-acetone virtually all of the injected Zn65 could be removed up to 1 hr after intravenous injection. However, extraction of the unbound zinc ion demonstrated that more than 90% of the Zn65 is bound to protein within the first 3 min after addition of Zn65. Dialysis of Zn65 against plasma protein showed the Zn65 ion is removed by plasma from solution by direct binding to nondialyzable plasma components. Irradiation of rats with supralethal dosages of X ray followed by subsequent injection of Zn65 demonstrated that clearance of Zn65 from plasma is unaffected by this major alteration of the metabolic state of the animal.

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