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Biomedical subjects

R Gräsbeck

Publications and source records attributed to R Gräsbeck.

At least 19 recordsLinked to original sources
PubMed1992

Preanalytical factors and standardized specimen collection: influence of psychological stress.

In order to devise and evaluate standardized specimen collection procedures, we studied the influence of psychological stress on the results of commonly analysed blood components: creatine kinase, lactate dehydrogenase, total protein and albumin in serum and blood picture. In addition, serum cortisol was assayed. Two kinds of stress were used: the Stroop test, a colour conflict task, and the thrill caused by the first jump of new parachutists. More changes were observed after the parachutist test than after the Stroop test. There was a difference in the responses of males and females. Females were more sensitive, especially to the parachutist test. Most of the changes observed were interpreted as being caused by haemoconcentration, possibly related to muscular tension. Cortisol, commonly used to indicate the level of stress, did not react much and is therefore not a good index of psychological stress.

Adolescent↗
PubMed1991

Binding assay and physicochemical characteristics of solubilized intrinsic factor receptor in ileal mucosal homogenates using phenyl-Sepharose to separate the saturated receptor from free intrinsic factor.

A radioisotopic assay was set to determine the physicochemical properties of the solubilized intrinsic factor receptor in pig mucosal extracts. In this assay, phenyl-Sepharose was used to separate the receptor-intrinsic factor-labelled cobalamin complex from the free saturated intrinsic factor. The association constant (at pH 7.4) of the receptor-intrinsic factor complex was estimated at 3.4 +/- 0.3 nM-1. Adsorption of the apo-receptor to phenyl-Sepharose allowed its binding site to be made accessible to intrinsic factor with an association constant in order of 6 nM-1. The receptor binding activity obtained with five mucosal extracts was closely correlated with that obtained by gel filtration of the intrinsic factor-receptor complex (r = 0.99). The radioisotope assay was used to detect the unsaturated receptor (apo-receptor) in sucrose density ultracentrifugation and in superose 6 gel filtration. The sedimentation coefficient was 9.5 s. The apo-receptor was eluted in three peaks in gel filtration, corresponding to the formation of oligomers. The peak of the monomer was increased in presence of EDTA. Its molecular mass was estimated at 270 kDa and its Stokes radius at 5.9 nm. It was concluded that calcium is involved in the oligomerisation of the apo-receptor.

Adsorption↗
PubMed1991

Studies on the transcobalamin receptor in hog kidney.

The binding of the cobalamin-transcobalamin complex by its solubilized receptor from hog kidney membrane was studied. The receptor bound the complex in a system containing bivalent cations, and the affinity was dependent on the NaCl concentration but not on temperature. The binding of cobalamin-transcobalamin to the receptor had an association constant of approximately 4.6 x 10(9) liter/mol and it was saturable and highly specific as competition by other proteins was not observed. The receptor had higher affinity for the cobalamin-transcobalamin complex (holo-TC) than for transcobalamin (apo-TC). Basic amino compounds known to interfere with tubular reabsorption of proteins did not inhibit the binding. Studies on subcellular fractions supported the view that the receptor was located on the brush border membrane of the kidney.

Animals↗
PubMed1990

Recommendation for collection of venous blood from children, with special reference to production reference values.

Reference values should be produced under standardized conditions. To enable comparison it is desirable to use the same procedure also in other clinical situations. A procedure for the collection of venous blood from children with special reference to production of reference values is recommended. It deals with five items: preparation of the child before specimen collection, preparation of the blood collection site, equipment for specimen collection, the specimen collection itself, and handling and storage of the specimen. Alternative methods are described since no single method is suitable for all paediatric age groups. The problem of adhering to a proposed procedure during routine clinical work is also commented upon. The recommendation has been produced as a joint effort of the Scandinavian Committee on Reference Values and a working group set up by the National Paediatric Societies in the Nordic countries.

Adolescent↗
PubMed1990

Reference values, why and how.

A review describing the fallacies of the concept of normal values and the reasons for its substitution with reference values. The "reference value philosophy", its terminology and recommended practical procedures are presented and it is warned not to use incorrect terms such as "normal reference range". The concepts of health and disease are discussed from the point of view of the laboratory. Reference values are not always derived from "healthy" persons. In order to be comparable, reference values and observed (i.e. patient) values should be produced in the same way, using the same analytical procedure, quality control, etc. The influence of preanalytical factors such as food intake, posture, use of tourniquet and freezing and storing samples is great and necessitates standardisation of specimen collection. Strategies for the selection of reference individuals are presented and the reasons and methods for subdivision of the data are described. Descriptions are also given of the individual reference values and the statistical treatment of collected data including multivariate analysis. Finally, the reporting of observed values in relation to reference values is discussed. There is a trend to avoid the use of the reference interval because of the temptation to regard the values falling outside it as pathological.

Data Collection↗
PubMed1989

Purification of intrinsic factor receptor from pig ileum using as affinity medium human intrinsic factor covalently bound to Sepharose.

Intrinsic factor receptor was purified from hog ileum using human intrinsic factor covalently bound to Sepharose. A yield of 49.6% and a specific activity of about 2500 pmol/mg protein were achieved. The purified receptor was very unstable: 24 h of storage or addition of sodium phosphate precipitated it. The association constant of the receptor for the cyano[57Co]cobalamin-intrinsic factor complex was estimated to be 2.1 nM-1. In native polyacrylamide gel electrophoresis it resolved in two 256 and 320 kDa bands; beta-mercaptoethanol treatment cleared it into four bands corresponding to molecular masses of 107, 81.8, 63.5 and 53.2 kDa. An additional 39.3 kDa band was considered to be an artefact due to the presence of Triton X-114. Isoelectric focusing polyacrylamide gel electrophoresis resolved the receptor into two isoproteins isoelectric at pH 4.7 and 5.1. A similar result was obtained in column electrofocusing with the 125I-iodinated receptor. The 125I-labelled receptor did not crossreact with rabbit anti-human intrinsic factor antiserum. The electrophoretic properties of the receptor purified with intrinsic factor covalently bound to Sepharose were compared to those of the receptor purified by the use of the classical cobalamin-affinity medium. It was concluded that a disassembled receptor was produced using the classical method.

Animals↗
PubMed1989

Lectin binding to the porcine and human ileal receptor of intrinsic factor-cobalamin.

The purified porcine receptor for the intrinsic factor-cobalamin complex bound to concanavalin A, lentil lectin and wheat germ lectin covalently coupled to Sepharose and was eluted with the corresponding soluble sugars. In contrast, human intrinsic factor bound efficiently to concanavalin A, to some extent to lentil lectin, but only slightly to wheat germ agglutinin. The binding of IF-Cbl to the receptor was inhibited when the receptor was pre-incubated with soluble wheat germ agglutinin, with an inhibition constant estimated to be 1.9 mumol/l. After transfer of the purified receptor from SDS-PAGE to Immobilon, ligand blotting of the purified receptor with iodinated lectin showed that concanavalin A and lentil lectin bound to three (75, 56 and 43 kDa) components but that wheat germ agglutinin bound only to the 75 kDa component. These results showed that the alpha subunit of the receptor could bind to wheat germ agglutinin, resulting in an inhibition of its binding with intrinsic factor. Both binding sites of intrinsic factor and of wheat germ agglutinin could be located near to each other.

Animals↗
PubMed1989

Reference values.

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Blood Chemical Analysis↗
PubMed1987

A rosette receptor assay with haem-microbeads. Demonstration of a haem receptor on K562 cells.

A rosette assay was developed for the detection of haem receptor-bearing cells. Indicator particles were prepared by covalent binding of haem to acrylic microbeads. The new method was tested on K562 human erythroleukaemia cells, known to take up haem. In tests on several batches, 80-90% of the K562 cells were rosetted with haem-microbeads whereas mature erythrocytes were haem receptor-negative. Rosette formation was inhibited in a dose-dependent manner by micromolar concentrations of free haemin but not by albumin or transferrin. Uncoated microbeads or albumin-coated microbeads did not attach to K562 cells but transferrin-coated microbeads rosetted 50-70% of them. Diferric transferrin inhibited these rosettes, but haemin had no effect.

Cell Line↗
PubMed1986

Isolation of the haemopexin-haem receptor from pig liver cells.

Isolated pig liver plasma membranes interact specifically with the haemopexin-haem complex (Kd 4.4 X 10(-7) M). Affinity chromatography was used to isolate a membrane component which binds this complex with high affinity. Pig serum haemopexin was first isolated by affinity chromatography on haemin-Sepharose followed by HPLC gel filtration. Liver membranes solubilized with Triton X-100 were incubated with haemin-Sepharose saturated with haemopexin, and as a control, with affinity gel lacking haemopexin. SDS-poly-acrylamide gel electrophoresis of the eluted protein indicated that from the haemin-Sepharose emerglow-molecular-mass haemin-binding proteins whereas the eluate from haemopexin-haemin-Sepharose contained an additional 71 kDa protein, which did not bind free haemin. This protein appears to represent the haemopexin-haem receptor or a part of it. Haem from the haemopexin complex, as also free haemin, was accepted by a binder in the plasma membrane, which in gel filtration behaved like an 80 kDa molecule. This component probably represents a second functional subunit of the haemopexin-haem receptor.

Animals↗
PubMed1979

An intestinal receptor for heme.

Porcine upper small intestine took up radioactive heme in vitro. The binder was localised to the brush borders, was solubilized with Triton X-100 and bound heme with high affinity. The binding was inhibited with non-radioactive heme but not with FeSO4, and control experiments indicated that it was not any of the known heme binders or aggregated heme. The binding factor could also be solubilized from human duodenum.

Animals↗
PubMed1979

Solubilization and preliminary characterization of the human ileal vitamin B12-intrinsic factor receptor.

The human ileal intrinsic factor receptor was solubilized with Triton X-100 using an improved method originally devised for the porcine receptor. At pH 7.4 and in the presence of Ca2+ the receptor bound the vitamin B12 complexes of normal human and pig intrinsic factor but not that of an abnormal biologically inert human intrinsic factor. EGTA dissociated vitamin B12-intrinsic factor from the receptor complexes. The solubilized vitamin B12-intrinsic factor receptor complex consisted of three to four molecular species termed HC-L, HC-20 S, HC-12 S and HC-8.5 S (the three last-mentioned referring to sedimentation coefficients). Of these HC-20 S was the dominating component and had a Stokes radius of 18 nm. Radioactive calcium was shown to be bound to vitamin B12-intrinsic factor and to be contained in its complex with the receptor.

Animals↗
PubMed1978

Comparative studies on intrinsic factor and cobalophilin in different parts of the gastrointestinal tract of the pig.

The vitamin B(12) binders in the pig pyloric mucosa gastric and intestinal juice from the upper gastrointestinal tract were fractionated into only two molecular forms, classified as intrinsic factor and cobalophilin. The unsaturated vitamin B(12)-binding power due to cobalophilin was lower in the intestinal than in the gastric juice. Electrofocusing revealed that intrinsic factor and cobalophilin in the intestinal juice contained more of the ;neutral'-type isoproteins, and the suggestion is made that this is due to enzyme activity. The gastric-juice intrinsic factor contained more acidic isoproteins, which supports the hypothesis that carbohydrate is added on to the polypeptide chain of this protein before it is secreted into gastric juice. The gastric- and intestinal-juice cobalophilins, studied also by electrofocusing, differed from that of pyloric mucosa and they appeared to be of salivary origin. With regard to molecular dimensions there was no significant difference between the intrinsic factors and cobalophilins from all sources studied. All cobalophilins had molecular weights by the formula of Svedberg of approx. 92500, Stokes radii of 4.62nm and sedimentation coefficients of 5.15S. The corresponding values for the intrinsic factors were 63600, 3.57nm and 4.38S. In addition, the intrinsic factors exhibited similar avidities for binding to the solubilized ileal intrinsic-factor receptor. Also the intrinsic factors and cobalophilins, irrespective of their source, bound to the analogous specific xenoantibodies with the same avidity. The present results demonstrate that intrinsic factor remains practically unaltered during its passage through the proximal intestine and render unlikely the speculations made about the presence of an endogenous binder for intrinsic factor as well as the existence of a ;pancreatic intrinsic factor'. In addition, they are compatible with the theory that the interference by undegraded cobalophilin may be the reason for the abnormal vitamin B(12) absorption observed in patients with pancreatic insufficiency.

Animals↗