Fate of receptor-bound human chorionic gonadotrophin in pseudopregnant rat ovaries perfused in vitro

1985 ◽  
Vol 109 (1) ◽  
pp. 115-121
Author(s):  
Hannu Rajaniemi ◽  
Jan Sogn ◽  
Paul Holmes ◽  
Björn Källfelt ◽  
Per Olof Janson
Keyword(s):  
Molecular Weight ◽  
Histological Examination ◽  
Gel Filtration ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Cell Periphery ◽  
Small Molecular Weight ◽  
Immunohistochemical Studies

Abstract. Pseudopregnant rats were injected with [125I] hCG, anaesthesized 1 h later and after cannulation of the aorta the ovaries were isolated and perfused with Gey & Gey buffer containing 0.2% BSA. The release of radioactivity was monitored for 2 h and analyzed by gel filtration. Five to ten per cent of the radioactivity was released within 2 h and represented small molecular weight peptides and iodotyrosine and [125I]hCG. Analysis of the ovarian radioactivity prior to and after perfusion revealed that virtually all hCG was receptor-bound. Loading the medium with unlabelled hCG displaced [125I]hCG from the receptor but did not enhance its degradation. Histological examination showed that the ovarian tissues were still intact after the 2 h perfusion. Immunohistochemical studies revealed a localization of the hCG at the cell periphery both prior to and after perfusion. These results provide evidence showing that the rate of internalization of receptor-hCG complexes in rat luteal cells is slow in vivo.

HEMODIALYSIS OF THE RAT

1966 ◽  
Vol 44 (5) ◽  
pp. 849-859 ◽  
Author(s):  
Sumner M. Robinson ◽  
David A. Hurwitz ◽  
Robert Louis-Ferdinand ◽  
William F. Blatt
Keyword(s):  
Molecular Weight ◽  
Gel Filtration ◽  
Low Molecular Weight

A technique is described for hemodialysis of either anesthetized or non-restrained rats. In the apparatus the dialysis plates of an autoanalyzer system are used with only minor modification. The efficiency of this method has been evaluated with regard to the clearance of saccharides, both in vitro and in vivo, as well as the extraction of nitrogenous low molecular weight moieties from circulating blood. Approximately 50% of the dialyzable material was obtained in a 1-hour dialysis. Further fractionation of the dialyzate was accomplished by gel filtration (Sephadex G-25).

scholarly journals Isolation and Characterization of Two Proteins fromMoraxella catarrhalis That Bear a Common Epitope

1998 ◽  
Vol 66 (9) ◽  
pp. 4374-4381 ◽  
Author(s):  
John C. McMichael ◽  
Michael J. Fiske ◽  
Ross A. Fredenburg ◽  
Deb N. Chakravarti ◽  
Karl R. VanDerMeid ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Bacterial Attachment ◽  
Vaccine Candidates ◽  
Isolation And Characterization ◽  
Sds Page

ABSTRACT The UspA1 and UspA2 proteins of Moraxella catarrhalisare potential vaccine candidates for preventing disease caused by this organism. We have characterized both proteins and evaluated their vaccine potential using both in vitro and in vivo assays. Both proteins were purified from the O35E isolate by Triton X-100 extraction, followed by ion-exchange and hydroxyapatite chromatography. Analysis of the sequences of internal peptides, prepared by enzymatic and chemical cleavage of the proteins, revealed that UspA1 and UspA2 exhibited distinct structural differences but shared a common sequence including an epitope recognized by the monoclonal antibody 17C7. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), purified UspA1 exhibited a molecular weight of approximately 350,000 when unheated and a molecular weight of 100,000 after being heated for 10 min at 100°C. In contrast, purified UspA2 exhibited an apparent molecular weight of 240,000 by SDS-PAGE that did not change with the length of time of heating. Their sizes as determined by gel filtration were 1,150,000 and 830,000 for UspA1 and UspA2, respectively. Preliminary results indicate the proteins have separate functions in bacterial pathogenesis. Purified UspA1 was found to bind HEp-2 cells, and sera against UspA1, but not against UspA2, blocked binding of the O35E isolate to the HEp-2 cells. UspA1 also bound fibronectin and appears to have a role in bacterial attachment. Purified UspA2, however, did not bind fibronectin but had an affinity for vitronectin. Both proteins elicited bactericidal antibodies in mice to homologous and heterologous disease isolates. Finally, mice immunized with each of the proteins, followed by pulmonary challenge with either the homologous or a heterologous isolate, cleared the bacteria more rapidly than mock-immunized mice. These results suggest that UspA1 and UspA2 serve different virulence functions and that both are promising vaccine candidates.

scholarly journals Characterization of proteins from the cytoskeleton of Giardia lamblia

1983 ◽  
Vol 59 (1) ◽  
pp. 81-103 ◽  
Author(s):  
R. Crossley ◽  
D.V. Holberton
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Giardia Lamblia ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Molecular Size ◽  
Molecular Weights

Proteins from the axonemes and disc cytoskeleton of Giardia lamblia have been examined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis. In addition to tubulin and the 30 X 10(3) molecular weight disc protein, at least 18 minor components copurify with the two major proteins in Triton-insoluble structures. The most prominent minor bands have the apparent molecular weights of 110 X 10(3), 95 X 10(3) and 81 X 10(3). Protein of 30 X 10(3) molecular weight accounts for about 20% of organelle protein on gels. In continuous 25 mM-Tris-glycine buffer it migrates mostly as a close-spaced doublet of polypeptides, which are here given the name giardins. Giardia tubulin and giardin have been purified by gel filtration chromatography in the presence of sodium dodecyl sulphate. Well-separated fractions were obtained that could be further characterized. Both proteins are heterogeneous when examined by isoelectric focusing. Five tubulin chains were detected by PAGE Blue 83 dye-binding after focusing in a broad-range ampholyte gel. Giardin is slightly less acidic than tubulin. On gels it splits into four major and four minor chains with isoelectric points in the pI range from 5.8 to 6.2. The amino acid composition of the giardin fraction has been determined, and compared to Giardia tubulin and a rat brain tubulin standard. Giardins are rich in helix-forming residues, particularly leucine. They have a low content of proline and glycine; therefore they may have extensive alpha-helical regions and be rod-shaped. As integral proteins of disc microribbons, giardins in vivo associate closely with tubulin. The properties of giardins indicate that in a number of respects - molecular size, charge, stoichiometry - their structural interaction with tubulin assemblies will be different from other tubulin-accessory protein copolymers studied in vitro.

In Vitro and in Vivo Antigen-Induced Release of High-Molecular Weight Neutrophil Chemotactic Activity from Human Nasal Tissue

1988 ◽  
Vol 2 (1) ◽  
pp. 7-11 ◽  
Author(s):  
T. Nagakura ◽  
T. Onda ◽  
Y. likura ◽  
T. Endo ◽  
H. Nagakura ◽  
...  
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Nasal Polyps ◽  
Gel Filtration ◽  
Chemotactic Activity ◽  
Nasal Tissue ◽  
Nasal Secretions ◽  
Neutrophil Chemotactic Activity

High molecular weight neutrophil chemotactic activity has been identified in resected human nasal polyps, inferior turbinates, and nasal secretions following antigen challenge. The estimated molecular weight, by gel filtration chromatography, was approximately 600,000. However, a heterogeneity of molecular weight in some patients was recognized. Our results suggest a possible role for high molecular weight-neutrophil chemotactic activity in the pathogenesis of hypersensitivity in the human nasal cavity.

STUDIES ON A HUMAN CHORIONIC GONADOTROPHIN-LIKE MATERIAL PRESENT IN NON-PREGNANT SUBJECTS

1978 ◽  
Vol 89 (3) ◽  
pp. 492-505 ◽  
Author(s):  
D. M. Robertson ◽  
H. Suginami ◽  
H. Hernandez Montes ◽  
C. P. Puri ◽  
S. K. Choi ◽  
...  
Keyword(s):  
Gel Filtration ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Specific Sequence ◽  
Gonadotrophin Secretion ◽  
Carboxy Terminal Region ◽  
Assay Methods ◽  
Low Levels ◽  
Carboxy Terminal

ABSTRACT The presence of an hCG-like material in urinary and pituitary extracts and plasma obtained from non-pregnant subjects was investigated. Two assay methods were used to detect this material following fractionation of pituitary and urinary extracts by gel filtration (Ultrogel AcA 54) and/or isoelectrofocusing: a) a radioimmunoassay employing an antiserum raised against a specific sequence of the carboxy terminal region (residues 115– 145) of the β-subunit of hCG, and b) an in vitro bioassay method which measures both hLH and hCG activities. The fractionation procedures employed provide a satisfactory separation of highly purified hCG and hLH preparations. In the pituitary and urinary extracts hCGβ-peptide-like immunoactive (PIA) material was found consistently, which co-eluted with iodinated hCG following gel filtration and possessed pI values similar to those of hCG when subjected to isoelectrofocusing. The PIA material also exhibited in vitro biological activity similar to that shown by hLH and hCG. Detectable levels of immunoactive material were also found in plasma; however, the plasma levels of this PIA material were not influenced by classical endocrine measures such as the stimulation or inhibition of gonadotrophin secretion. The low levels of this material in plasma precluded its further characterization by gel filtration or electrofocusing. Whereas the present data and those reported by other investigators seem to suggest the presence of some hCG-like material in urinary and pituitary extracts and possibly in plasma of non-pregnant subjects, it is emphasized that the available evidence is not sufficiently conclusive to exclude other interpretations as to the nature of this material.

Immunochemical analysis of the human chorionic gonadotrophin-like material secreted by 'normal' and neoplastic urothelial cells

1989 ◽  
Vol 2 (2) ◽  
pp. 107-112 ◽  
Author(s):  
R. K. Iles ◽  
T. Chard
Keyword(s):  
Bladder Tumour ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Sds Page ◽  
Β Hcg ◽  
Bladder Carcinomas ◽  
Immunochemical Characteristics ◽  
Normal Urothelium

ABSTRACT Material with the immunochemical characteristics of human chorionic gonadotrophin (hCG) is produced by bladder tumour cells in vitro and in vivo. In order to characterize this material further, media were collected from 17 cell cultures (three choriocarcinomas, seven bladder carcinomas and seven 'normal' urothelium). The hCG-like material was compared with pregnancy hCG and purified α- and β-subunits by specific radioimmunoassays. Media were also submitted to affinity chromatography and the fractions further analysed by SDS-PAGE and Western blotting. It was shown that both the neoplastic and normal urothelium produced only free β-subunit-like material. This urothelial 'β-hCG' has the same molecular weight and electrophoretic mobility as that present in the intact hCG of pregnancy.

INTERNATIONAL REFERENCE PREPARATION OF HUMAN CHORIONIC GONADOTROPHIN FOR IMMUNOASSAY: POTENCY ESTIMATES IN VARIOUS BIOASSAY AND PROTEIN BINDING ASSAY SYSTEMS; AND INTERNATIONAL REFERENCE PREPARATIONS OF THE α AND β SUBUNITS OF HUMAN CHORIONIC GONADOTROPHIN FOR IMMUNOASSAY

1980 ◽  
Vol 84 (2) ◽  
pp. 295-310 ◽  
Author(s):  
P. L. STORRING ◽  
ROSE E. GAINES-DAS ◽  
D. R. BANGHAM
Keyword(s):  
Geometric Mean ◽  
Collaborative Study ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Considerable Heterogeneity ◽  
International Reference ◽  
Reference Preparation ◽  
Α And Β Subunits

The preparation and nature of the International Reference Preparation of Human Chorionic Gonadotrophin (HCG) for Immunoassay (IRP), as well as that of a second batch of ampoules (HCG 75/589) prepared identically from the same HCG preparation, are described. A collaborative study of these materials was carried out by 11 laboratories in eight countries, using different bioassay and immunoassay methods. Using the various in-vivo and in-vitro bioassays and receptor assays, the mean log potency estimates for each method within each laboratory of the HCG content of ampoules of the IRP, in terms of the Second International Standard of Human Chorionic Gonadotrophin for Bioassay (IS), were homogeneous and gave an overall weighted geometric mean (95% confidence limits) of 650 (632–669) International Units (i.u.)/ampoule. There was considerable heterogeneity of potency estimates of the IRP in terms of the IS both within and between many of the immunoassay systems (reflecting the impurity of the IS), and hence attempts to calibrate the IRP with immunoassay systems of different specificities were invalid. Immunoassay estimates of the HCG content of preparations of serum and urine, in terms of the IRP, showed considerable heterogeneity between assay systems (although the degree of this heterogeneity was no greater than that observed using the IS as standard), but the ranking order between preparations was consistent. Confirmation was obtained that contamination of the IRP with HCG-α and HCG-β subunits was insignificant. Accelerated degradation studies of the IRP stored at increased temperatures suggested that its stability under normal storage conditions would be satisfactory. It was agreed that the IRP was suitable to serve as an international reference preparation for immunoassay, and it was assigned a unitage of 650 i.u./ampoule on the basis of bioassay calibration. Since the ampoules of HCG (75/589) did not differ significantly from the IRP in any of the assay systems studied, it appeared to be equally suitable as a reference preparation. The International Reference Preparations of the α and β Subunits of Human Chorionic Gonadotrophin for Immunoassay are also described.

ÉTUDE DE L'ACTIVITÉ BIOLOGIQUE IN VITRO DES HORMONES GONADOTROPES

1963 ◽  
Vol 42 (4) ◽  
pp. 509-513 ◽  
Author(s):  
D. Gospodarowicz ◽  
J. Legault-Démare
Keyword(s):  
Incubation Medium ◽  
Cholesterol Synthesis ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Corpora Lutea ◽  
Lactogenic Hormone ◽  
Normal Rat ◽  
Stimulation Of

ABSTRACT Human chorionic gonadotrophin (HCG) and lactogenic hormone (LTH or prolactin) were found practically inactive on the incorporation of 14Cacetate into cholesterol of normal rat corpus luteum in vitro. On the contrary, when added simultaneously to the incubation medium, they increased by 90% the labeling of cholesterol. When pseudopregnancy corpora lutea were used, HCG alone stimulated to the same amount, but no stimulation was observed with LTH alone. These results show that the stimulation of cholesterol synthesis is produced by a synergic action of LTH and HCG, LTH being introduced either in vivo (pseudopregnancy) or in vitro.

Iron availability from meat

1978 ◽  
Vol 39 (3) ◽  
pp. 631-638 ◽  
Author(s):  
T. Hazell ◽  
D. A. Ledward ◽  
R. J. Neale
Keyword(s):  
Molecular Weight ◽  
Degradation Products ◽  
Gel Filtration ◽  
Rat Muscle ◽  
Muscle Extract ◽  
Freeze Dried ◽  
Absorption Studies ◽  
Fe Complexes

1. The distribution of radioactive iron in59Fe-labelled rat muscle extract was determined using ge filtration. This showed that most (approximately 70%) of the radioactivity was associated with the heamatin compounds; myoglobin and haemoglobin.2. Raw beef and freeze-dried rat muscle were digested in vitro, under simulated physiological conditions, and after centrifugation the supernatants fractionated by gel filtration. The soluble products were haematin Fe complexes of molecular weight above 10 000 and non-haematin Fe compounds of molecular weight below 6000, the major products being the non-haematin Fe complexes. The soluble compounds were also separated by dialysis and, in rat muscle, it was found that the low-molecular-weight non-haematin compounds accounted for more than 80% of the total soluble iron.3. In vivo absorption studies with rats showed the Fe in a digested muscle dialysate to be more readily absorbed than that from an aqueous muscle extract which itself was more readily absorbed than the Fe from whole blood.4. It may not, therefore, be the haemoproteins per se which are responsible for the high availability of Fe in meat, but rather the nature of their degradation products, formed by digestion within the meat environment.

Sign in / Sign up

Export Citation Format

Share Document