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1.
Monoclonal antibody specific for yeast elongation factor 3 总被引:1,自引:0,他引:1
Hybridomas have been prepared by fusing mouse myeloma (P3 X 63 Ag8) cells with spleen cells of mice immunized with a yeast fraction enriched with respect to non-ribosomal translational components. Cloned hybridoma lines were grown in the form of ascites tumors, and the monoclonal antibodies produced were purified from the ascites fluid by chromatography on DEAE-Affi-Gel Blue. One of the antibodies, from a hybridoma cell line designated as PSH-1, inhibited the translation of natural mRNA and poly(U) and polysomal chain elongation in a cell-free protein-synthesizing system from yeast. Resolution and partial purification of the elongation factors indicated that the monoclonal antibody from PSH-1 did not interact with EF-1 or EF-2 but reacted with and inactivated EF-3, the 125 000 molecular weight additional elongation factor specifically required with yeast ribosomes. The EF-3 purified from the cytosol by immunoaffinity chromatography was comparable to that prepared by ion-exchange chromatography. Evidence was obtained which indicated that EF-3 was essential for the translation of natural mRNA as well as poly(U), was associated with polysomes but not ribosomal subunits, and was required for every cycle in the elongation phase of protein synthesis. 相似文献
2.
Monoclonal antibody specific for the T-tubule of skeletal muscle 总被引:1,自引:0,他引:1
N N Malouf S Taylor G Y Gillespie J M Bynum P E Wilson G Meissner 《The journal of histochemistry and cytochemistry》1986,34(3):347-355
Monoclonal antibodies were raised against a triad-enriched (sarcoplasmic reticulum-T-tubule complex) microsomal membrane fraction of rabbit skeletal muscle. The avidin-biotin complex (ABC) immunoperoxidase staining method was used to screen hybrid colonies. Positive antibodies exhibited a granular doublet pattern at the A-I junction, consistent with the location of triads in rabbit muscle. One monoclonal antibody, M171, was further characterized by ultrastructural and immunoadsorption techniques. Postembedding electron immunocytochemistry was performed on tissue sections embedded in Lowicryl K4M. Goat anti-mouse immunoglobulin absorbed to 10 nm colloidal gold particles was used as an ultrastructural label. In these studies, M171 recognized an epitope at the triads and at periodic openings along the plasmalemma. Immunoadsorption on protein transfers of isolated sarcoplasmic reticulum, surface membrane (plasmalemma and T-tubule), and triad-enriched fractions showed that M171 reacts with a surface membrane component. Taken together, these studies suggest that M171 recognizes an epitope associated with the T-tubule at the triad and at the "mouth" of the T-system at the plasmalemma. 相似文献
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4.
T Kisaki T F Huff D H Conrad J Yodoi K Ishizaka 《Journal of immunology (Baltimore, Md. : 1950)》1987,138(10):3345-3351
A B cell hybridoma secreting monoclonal antibody against human IgE binding factors was obtained by immunization of BALB/c mice with partially purified IgE binding factors, and fusion of their spleen cells with SP-2/0-AG14 cells. The monoclonal antibody bound all of the 60,000, 30,000, and 15,000 dalton IgE binding factors from two T cell hybridomas and those from activated T cells of a normal individual. The antibody bound both IgE-potentiating factors, which had affinity for lentil lectin, and IgE-suppressive factors, which had affinity for peanut agglutinin. However, the monoclonal anti-IgE-binding factor bound neither Fc epsilon R on RPMI 8866 cells nor IgE binding factors from the B lymphoblastoid cells. A monoclonal antibody against Fc epsilon R on B cells (H107) bound the 60,000 and 30,000 dalton IgE binding factors from both T cell hybridomas and RPMI 8866 cells but did not bind the 15,000 dalton IgE binding factors from either T cells or B cells. The results indicate that T cell-derived IgE binding factors have a unique antigenic determinant that is lacking in both Fc epsilon R on B cells and B cell-derived IgE binding factors. The anti-IgE binding factor and anti-Fc epsilon R monoclonal antibodies both failed to stain cell surface components of IgE binding factor-producing T cell hybridomas. However, both antibodies induced the T cell hybridoma to form IgE binding factors. The results suggest that the T cell hybridomas bear low numbers of Fc epsilon R that share antigenic determinants with IgE binding factors secreted from the cells. 相似文献
5.
Monoclonal antibody specific for calf thymus RNA polymerases IIO and IIA 总被引:12,自引:0,他引:12
6.
We describe here the development of a new hybridoma cell line, CF12F6, that produces a specific antibody to Botryllus schlosseri (a colonial tunicate). The monoclonal antibody was isotyped as IgG1 (by enzyme-linked immunosorbent assay), and the cellular localization of the antigenic epitope that reacts specifically with CF12F6 was confined to the cells of the pyloric gland of the zooid (by immunohistochemistry). The pyloric gland participates in osmoregulation, digestion, glycogen storage, and various other secretion functions that will be studied further by the use of monoclonal antibody CF12F6, the first in botryllid ascidians that recognizes cells of a whole, single organ. 相似文献
7.
F Hirose P Kedar T Takahashi A Matsukage 《Biochemical and biophysical research communications》1985,132(1):210-216
Four monoclonal antibodies against chicken DNA polymerase alpha were obtained from mouse hybridomas (see ref. 1). Two of them, 4-2D and 4-8H, recognized different epitopes of the DNA polymerase alpha-DNA primase complex as determined by a competitive enzyme-linked immunosorbent assay. Antibody 4-8H partially (about 30%) neutralized the combined activity of primase-DNA polymerase alpha as well as the DNA polymerase alpha activity. In contrast, antibody 4-2D did not neutralize DNA polymerase alpha activity, but neutralized the primase-DNA polymerase alpha activity extensively (up to 80%). Furthermore, although an immunoaffinity column made with 4-8H antibody retained virtually all of the DNA polymerase alpha with and without associated primase, a column made with 4-2D antibody did not bind DNA polymerase alpha without the primase, but retained the enzyme associated with the primase. These results indicate that 4-8H monoclonal antibody is specific for DNA polymerase alpha and 4-2D monoclonal antibody is specific for the primase or a special structure present in the primase-DNA polymerase alpha complex. 相似文献
8.
Removal of blood group anti-A and anti-B antibodies can prevent hyperacute organ rejection in ABO-incompatible transplantation. We are developing an extracorporeal-specific antibody filter (SAF) as an immunoadsorption device for direct removal of ABO blood group antibodies from whole blood, without the need for plasma separation and plasma exchange. A hollow fiber-based small scale SAF (mini-SAF) device was fabricated and synthetic A antigen, Atrisaccharide (Atri) conjugated to activated polyacrylic acid, was immobilized on the fiber lumen surface. Monoclonal antibody anti-A IgM were specifically removed up to 70% of initial antibodies using mini-SAF device. The monoclonal anti-A capture experiments on mini-SAF indicated that antibody removal relative to the initial concentration is independent of inlet concentration in the beginning; however, as the surface starts saturating with bound antibodies, removal becomes dependent on inlet concentration. No significant effect of flow rate on removal rate was observed. The radial diffusion and axial convection-based mathematical model developed for unsteady state antibody removal was in good agreement with the experimental data and showed that the antibody removal rate can be maximized by increasing the antibody-binding capacity of the SAF. 相似文献
9.
Monoclonal antibody specific for carbodiimide-fixed glutamate: immunocytochemical localization in the rat CNS 总被引:2,自引:0,他引:2
Glutamate is widely distributed in the central nervous system (CNS) and is present in greater amounts than any other putative neurotransmitter. To study its distribution in the CNS, a monoclonal antibody was raised against gamma-L-glutamyl-L-glutamic acid (gamma-Glu-Glu) conjugated to keyhole limpet hemocyanin (KLH) using glutaraldehydeborohydride. By use of this antibody, indirect immunoperoxidase staining was observed in CNS tissue fixed with carbodiimide to form gamma-Glu-Glu from glutamate and post-fixed with glutaraldehyde or paraformaldehyde. In contrast, immunoreactivity was quite low in tissues fixed only with glutaraldehyde. Absorption controls indicated that the staining of carbodiimide-fixed tissue could be inhibited by micromolar concentrations of gamma-Glu-Glu but not by other small molecules. Using ELISA, the antibody reacted strongly with the gamma-Glu-Glu/KLH conjugate used to immunize the mouse, but not with other small molecules conjugated to KLH. The reactivity of the antibody with the gamma-Glu-Glu/KLH conjugate on ELISA was inhibited by free gamma-Glu-Glu in micromolar concentrations, but not by similar dipeptides or amino acids. Dense immunocytochemical staining was observed in cortical pyramidal cells, cerebellar granule cells, and the cochlear nuclei. Staining with this monoclonal antibody correlated well with other methods of localizing glutamate in the CNS. 相似文献
10.
Monoclonal antibody specific for Listeria monocytogenes, Listeria innocua, and Listeria welshimeri 总被引:3,自引:0,他引:3
Eight hundred fifty-nine murine hybridomas were produced from eight fusions, and 27 were characterized for secretion of antibodies reactive to Listeria monocytogenes. One monoclonal antibody (MAb), P5C9, reacted with all test strains of L. monocytogenes (31 of 31), L. innocua (3 of 3), and L. welshimeri (1 of 1) but not with any strains of the other four Listeria species or with any of 22 gram-positive or 11 gram-negative species of bacteria when tested in microtiter and dot blot enzyme immunoassays. Of the other 26 antibodies, 20 reacted with either L. monocytogenes Scott A or V7 and with some or all of the other six Listeria species but also cross-reacted with some or all of the non-Listeria bacteria tested. MAb P5C9 is of the immunoglobulin G1 murine subclass. In Western blot (immunoblot) analyses, this MAb reacted with a single antigen with a molecular weight of 18,500, and it is shared in common with all three reactive species, L. monocytogenes, L. innocua, and L. welshimeri. This antigen was extracted with detergent and appeared to be cell bound. 相似文献
11.
Monoclonal antibody technology for mycotoxins 总被引:4,自引:0,他引:4
Specific monoclonal antibodies (MABs) against aflatoxins, ochratoxin A, zearalenone, diacetoxyscirpenol and T-2 toxin have been prepared in various laboratories by the application of hybridoma technology to mycotoxins. These antibodies can be selected for sensitivity, reduced cross-reactivity, reliability and ease of production. When a suitable antibody is chosen it can then be used in a rapid immunological method such as an enzyme-linked or radio-immunoassay or immunoaffinity chromatography system. These assays have a lower limit of mycotoxin detection in the ng/ml range and have been applied to the determination of mycotoxins in samples such as maize, peanuts, peanut butter, milk and porcine kidneys. Using these immunoassay techniques, sample preparation has generally been simplified to a matter of solvent extraction of mycotoxins from the sample followed by dilution; under these conditions, levels of 1-5ug of mycotoxins/kg of sample can be found. The application and advantages of MABs to mycotoxins and the use of these antibodies in various assay techniques is discussed. 相似文献
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A monoclonal antibody (MAb), EM-7G1, specific for Listeria monocytogenes was developed by using a previously developed MAb, C11E9 (A. K. Bhunia, P. H. Ball, A. T. Fuad, B. W. Kurz, J. W. Emerson, and M. G. Johnson, Infect. Immun. 59:3176-3184, 1991), to mask epitopes shared by L. monocytogenes and Listeria innocua in a 66-kDa cell surface protein. MAb EM-7G1 was an immunoglobulin subclass G1 antibody with kappa light chains. This MAb reacted with all 34 strains of L. monocytogenes tested and showed no cross-reaction with other Listeria spp. or other gram-positive or gram-negative organisms tested by enzyme-linked immunosorbent assay, dot blotting, and colony blotting. A second MAb, EM-6E11, reacted with all Listeria spp. tested but no other bacteria. In a Western blot (immunoblot) assay, EM-7G1 reacted with a crude cell surface protein of 66 kDa with a pI value of 6.7, while EM-6E11 reacted with two protein bands of 43 and 94 to 97 kDa with pI values of 4.0 and 4.3, respectively. Results with trypsin or pronase treatments indicated that the cell antigen reacting with EM-7G1 was on the surface of L. monocytogenes V7 and Scott A cells. 相似文献
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15.
Monoclonal antibodies specific for beta-adrenergic ligands 总被引:3,自引:0,他引:3
S Chamat J Hoebeke A D Strosberg 《Journal of immunology (Baltimore, Md. : 1950)》1984,133(3):1547-1552
After somatic cell fusion between splenocytes of immunized BALB/c mice and NS-1 myeloma cells, eight clones were obtained secreting anti-alprenolol antibodies as characterized by means of an ELISA. Four of these were subcloned and were studied further. The association constant for alprenolol ranged from 1.9 X 10(6) M-1 to 24 to 10(6) M-1. Competitive inhibition of [3H]-l-dihydroalprenolol binding revealed cross-reactivity with beta-adrenergic ligands, with a higher avidity for antagonists than for agonists. Two of the antibodies had a higher affinity for the l-isomer than for the d-isomer. The most stereospecific of these antibodies showed only affinity for beta-adrenergic antagonists and for the agonist isoproterenol. The other recognized both beta-adrenergic antagonists and agonists; it also showed an increase in tryptophan fluorescence after ligand binding. This property was used for the physicochemical study of the hapten-antibody interaction. 相似文献
16.
J A Sawchak J H Sher S A Shafiq 《The journal of histochemistry and cytochemistry》1989,37(10):1539-1543
We report the characterization of monoclonal antibody (MAb) ALD 180, prepared against the myosin of slow avian muscle, for studies of human muscle development and disease. With the use of radioimmunoassays, Western immunoblots of native and denatured myosins, and epifluorescent indirect immunocytochemistry, we show that ALD 180 is specific for an epitope in human prenatal skeletal muscle myosin heavy chain (MHC), which is expressed in diminishing abundance in fetal fibers from at least 19-22 weeks' gestation to term and also in regenerating muscle fibers seen in diseased muscles from both children and adults. ALD 180 recognizes an epitope apparently different from those reacting with anti-prenatal human myosin MAb previously described, and therefore affords a complementary reagent for use in future studies of human myosin isoform expression and regulation. 相似文献
17.
Coiffier B 《Comptes rendus biologies》2006,329(4):241-254
Rituximab was the first monoclonal antibody to have been registered for the treatment of B-cell lymphomas. Randomized studies have demonstrated its activity in follicular lymphoma, mantle-cell lymphoma, and diffuse large B-cell lymphoma in untreated or relapsing patients. Because of its high activity and low toxicity ratio, rituximab has transformed the outcome of patients with B-cell lymphoma. A combination of rituximab plus chemotherapy, R-CHOP, has the highest efficacy ever described with any chemotherapy in diffuse large B-cell lymphoma and follicular lymphoma. The role of radio-labelled antibodies is still to be defined. 相似文献
18.
Mycoplasma infection of cell cultures is still a major problem in some laboratories. Although several methods can be used for their detection, identification is normally by serological procedures. As no commercial source for the necessary antibodies is available we have prepared monoclonal antibodies to the five mycoplasma species that account for the majority of cell culture infections. These antibodies have been characterized by the growth inhibition test (GIT), immunofluorescence, and enzyme linked immunosorbent assay (ELISA) and have shown perfect correlation in all tests when compared to conventional antisera raised in rabbits or donkeys. In addition, a monoclonal antibody to Mycoplasma pneumoniae was produced. M. pneumoniae is an infrequent cell culture contaminant but is a human pathogen, and the monoclonal antibody described here could be useful in the clinical diagnosis of M. pneumoniae infection in man. 相似文献
19.
Monoclonal antibodies specific for cell culture mycoplasmas 总被引:3,自引:0,他引:3
David W. Buck Roger H. Kennett Gerard McGarrity 《In vitro cellular & developmental biology. Plant》1982,18(4):377-381
Summary Mycoplasma infection of cell cultures is still a major problem in some laboratories. Although several methods can be used
for their detection, identification is normally by serological procedures. As no commercial source for the necessary antibodies
is available we have prepared monoclonal antibodies to the five mycoplasma species that account for the majority of cell culture
infections. These antibodies have been characterized by the growth inhibition test (GIT), immunofluorescence, and enzyme linked
immunosorbent assay (ELISA) and have shown perfect correlation in all tests when compared to conventional antisera raised
in rabbits or donkeys. In addition, a monoclonal antibody toMycoplasma pneumoniae was produced.M. pneumoniae is an infrequent cell culture contaminant but is a human pathogen, and the monoclonal antibody described here could be useful
in the clinical diagnosis ofM. pneumoniae infection in man.
These studies were supported by Grants Al-15748 from the National Institute of Allergy and Infectious Diseases, and GM20138-07
from the National Institutes of Health, Bethesda, MD. 相似文献
20.
The IgG1hybridoma antibody, 91.9H, was originally raised against sulfated
mucins isolated from normal human colonic mucosa. Previous studies have
shown that the 91.9H antigen is expressed on normal colonic epithelial
cells and the sulfomucins that they produce, but not in the normal small
intestine and stomach. Tissue-specific changes occur in 91.9H antigen
expression in disease: the antigen diminishes in colonic carcinomas,
whereas in regions of gastric mucosa showing intestinal metaplasia and in
gastric carcinomas, the antigen is expressed as a "neo-antigen." This
report is concerned with elucidation, by the neoglycolipid technology, of
the determinant recognized by antibody 91.9H using sulfated and sialyl
oligosaccharides of Lewisa(Lea) and Lextypes, and analogs that lack
sulfate, sialic acid, or fucose. Binding experiments with the lipid-linked
oligosaccharides immobilized on chromatograms or on microwells, and
inhibition of binding experiments with free oligosaccharides based on di-,
tri- and tetrasaccharide backbones, show that the 91.9H antigenic
determinant is based on a trisaccharide backbone, and consists of the
3'-sulfated Leatetrasaccharide sequence, which is a potent ligand for the
E- and L-selectins. The antibody gives a relatively low signal with the
3'-sulfated non-fucosylated backbone, and has no detectable cross- reaction
with the 3'-sulfated Lexisomer, nor with sialyl-Leaand - Lexanalogues.
Antibody 91.9H is a valuable addition, therefore, to the repertoire of
reagents for mapping details of the distribution, and determining the
relative importance of sulfated and sialyl oligosaccharides as ligands for
the selectins, in normal and pathological epithelia and endothelia.
相似文献