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Biomedical subjects

R Wahl

Publications and source records attributed to R Wahl.

At least 91 records · Page 5Linked to original sources
PubMed1987

Molecular weight determination of allergen extracts and isolation of allergenic molecules by high-performance liquid chromatography.

The usefulness of size-exclusion high-performance liquid chromatography (HPLC) for the molecular weight determination of allergen extracts as a method of process control was investigated by measuring seven different batches of timothy pollen extracts. Preparative HPLC was used for the isolation of the major allergen Cat 1, from a crude cat hair and skin scraping (CHSS) extract. The chromatograms of the seven batches of the timothy pollen extracts looked very similar, suggesting the same molecular weight distribution in each extract. The major allergen, Cat 1, could be isolated from the crude CHSS extract by preparative HPLC in acceptable purity, determined by crossed radioimmunoelectrophoresis.

Allergens↗
PubMed1987

Chronic cardiac reactions. IV. Effect of drugs and altered functional loads on cardiac energetics as inferred from myofibrillar ATPase and the myosin isoenzyme population.

A major determinant of myocardial energetics is the ATPase activity of myofibrils. In order to account for chronic changes in myofibrillar ATPase, the state equation of the intertropomyosin-interaction model of Tawada et al. was extended by introducing the rates of cross-bridge cycling of myofibrils composed of V-1 or V-3 and the concentration of the myosin isoenzymes. Cross-bridge cycling rates of 1.0 or 0.7 were derived for myofibrils composed of V-1 or V-3, respectively. Ca2+ responsiveness and positive co-operativity were not significantly affected by the myosin isoenzymes. Redistribution of the myosin isoenzyme population and thus altered myocardial energetics was observed following administration of various drugs and as a result of different functional loads. Besides thyroid hormones, catecholamines had a marked influence on myosin. Reducing the adrenergic drive by administration of atenolol, guanethidine or reserpine led to a shift in the direction of V-3. Since serum T3 levels were not significantly reduced by these interventions, the drugs act most probably at the organ level. The functional states responsible for the increase in the proportion of V-3 (pressure load, intermittent feeding, schedule-induced stress) also did not affect circulating T3 in a manner that could entirely explain the redistribution. Hypertrophy-induced dilution of sympathetic nerve fibres or reduced adrenergic responsiveness most likely play a role in the redistribution. An increase in the proportion of V-1 was observed following swimming exercise but not, however, after spontaneous or enforced running. In the swim-exercised rats, T3 was markedly reduced. Thus, the trigger reactions linked most probably to the high adrenergic drive during swimming have to overcome the lower T3 level. It is concluded that myocardial energetics can be decisively altered by a variety of drugs and functional loads, whereby the trigger reactions leading to an altered gene expression of myosin cannot be accounted for entirely by altered circulating T3 but most probably involve the adrenergic system.

Adenosine Triphosphatases↗
PubMed1987

Monoclonal antibody (5G6.4) against ovarian carcinoma shows inhibition of in vitro colony formation.

Monoclonal antibodies (MAbs) have the potential for diagnosis and therapy of cancer, 5G6.4 is a MAb of the IgG2a class which was produced by immunization of BALB/c mice with human ovarian carcinoma (Ov Ca) cells. To further characterize 5G6.4, its effect on cell growth was tested using a human Ov Ca cell line established in our laboratory. A clonogenic assay was set up in 48-well plates in a double agar system. The cells were plated and 5G6.4 was added at different concentrations. Control plates consisted of cells with media without MAb. Negative control plates were also prepared using the same concentrations of an isotype-matched antimelanoma MAb, 225.28s. Colony formation (CF) was reduced to 50% or less of control with increasing amounts of 5G6.4 up to 50 micrograms/ml. Although CF was still depressed at concentrations above 50 micrograms/ml, the inhibition did not follow a directly proportional line; instead, it followed a bell-shaped curve. Plates with the control MAb, 225.28s, did not show this response. Similar results were obtained with cells from malignant Ov Ca ascites in the same clonogenic assay. Our study suggests that in the evaluation of the in vitro effect of MAb on growth, the concentration of MAb is crucial and may not show a linear response and that 5G6.4 may have a direct therapeutic effect by blocking the growth of Ov Ca cells. 5G6.4 is presently under study for therapy in an animal model.

Antibodies, Monoclonal↗
PubMed1987

Products of cells cultured from gliomas. V. Cytology and morphometry of two cell types cultured from glioma.

Explants of cells of a human glioma were evaluated with the nuclear fluorochrome 4',6-diamidino-2-phenylindole, by phase-contrast illumination, and by Giemsa staining correlated with double immunofluorescence for glial fibrillary acidic protein (GFAP) and fibronectin (FN). FN-positive (FN+) cells lacked GFAP detectable by immunofluorescence. Their mean nuclear-to-cytoplasmic ratio was large (0.192). Actual mean areas of nuclei (1,252 microns2) and cytoplasm (8,376 microns2) of FN+ cells compared with mean areas of fibroblasts suggested that the high nuclear-to-cytoplasmic ratio of FN+ cells was due to their microscopically evident reduced cytoplasmic spreading rather than to larger nuclei. Some FN+ cells showed marked variation in nuclear and nucleolar size and shape. Others had abnormal mitoses or hyperchromatic nuclei. GFAP-positive (GFAP+) cells lacked FN detectable by immunofluorescence. GFAP+ cells were smaller and less round than FN+ cells. Their usual location was growing on a layer of FN+ cells. The mean nuclear-to-cytoplasmic ratio (0.245) of GFAP+ cells was the highest in the study, surpassing the ratio of the continuous glioma line LM (0.176). Mean areas of nuclei (289 microns2) and of cytoplasm (1,350 microns2) of GFAP+ cells suggested that their high nuclear-to-cytoplasmic ratio was due to their microscopically evident reduced cytoplasmic spreading. Reduced spreading was associated with extension of long, thin cytoplasmic processes. The majority of GFAP+ cells showed marked cytoplasmic basophilia, nuclear hyperchromasia, and clumped chromatin. Features observed in both FN+ and GFAP+ cells from this high-grade astrocytoma are features associated with malignant transformation in more thoroughly studied tumor systems.

Antigens, Neoplasm↗
PubMed1987

Application of the first international standard of Dermatophagoides pteronyssinus (house dust mite) in the evaluation of allergen extracts produced from two different source materials.

The first international standard (IS) of the house dust mite Dermatophagoides pteronyssinus was used to compare side-by-side two allergen extracts prepared from two different house dust mite sources, namely whole mite cultures (WMC) and purified mite bodies (PMB), by employing several biochemical and immunochemical methods. In most methods employed, IS and WMC resembled more to each other than to PMB. These similarities comprehend protein contents, protein patterns, antigen and allergen patterns as well as total in vitro activity obtained in RAST inhibition (RI). The differences between WMC and PMB could be reproduced in different batches. Standardization in relation to IS of D. pteronyssinus yields activity ratios ranging from 2.4 to 70.1 between WMC and PMB depending on the variation of RI employed, indicating quantitative differences between the two preparations. When coupled to BrCN activated paper disks and employed in direct RAST, both preparations yielded almost identical binding values of specific IgE. Also in skin prick test, WMC and PMB produced very similar results in 12 mite-allergic patients. It was not possible to differentiate between the two preparations when used in concentrations which are usually employed for diagnostic purposes. These findings demonstrate the diversity of allergens in mite extracts derived from different sources and reveal the problems which are involved with the use of the WHO D. pteronyssinus IS.

Allergens↗
PubMed1987

Allergenic activity of short ragweed pollen (Ambrosia elatior) from different years and/or suppliers: criteria for the selection of an in-house allergen reference preparation.

We have investigated dialyzed extracts (A-F) of short ragweed pollens (Ambrosia elatior) from six consecutive production years and four different companies by protein nitrogen units, high performance liquid chromatography, isoelectric focusing, SDS-PAGE, RAST inhibition, crossed radioimmunoelectrophoresis, and antigen E contents. Extract A showed lowest yields in all quantitative assays. Isoelectric focusing, SDS-PAGE, crossed immunoelectrophoresis, and crossed radioimmunoelectrophoresis showed comparable patterns for all extracts except for extract A. Apparently, the activity of short ragweed pollens depends on the year of production and/or the skill of the producer. These findings are in contrast to similar examinations of other pollen species.

Allergens↗
PubMed1987

A manufacturer's criteria for in-house reference preparations for RAST inhibition.

Reagents employed for RAST-inhibition need careful characterization. An allergen reference preparation was produced from carefully selected raw materials. After immunochemical and biochemical characterization, the reference preparation was used to produce allergen reference paper disks. Their quality was controlled by the amount of bound protein and serum titration, which should yield an S-shaped dose-response curve and specific binding of IgE of at least ten times more than background binding. In parallel, the biologic activity of the allergen reference preparations was established in quantitative skin prick tests in at least 20 allergic patients. From these patients, serum samples were collected and pooled after careful characterization. All three necessary reagents for RAST-inhibition (extract, disks, and serum pool) were stabilized by freeze-drying. The reproducibility of the assay procedure itself was checked by documenting the 50% inhibition value of the reference preparation, accepting only values within +/- 2 SD. By employing such reagents we could measure the activity of alum-adsorbed grass pollen extracts with sufficient precision, ie, the coefficient of variation of the mean potency of 12 batches was less than 40%.

Allergens↗
PubMed1986

High-performance liquid chromatographic molecular weight determination of allergen extracts. Examination of the influence of the column material on allergenic activity and allergen patterns.

To investigate whether the column material employed in size-exclusion high-performance liquid chromatography (HPLC) influences the allergenic activity and antigen/allergen patterns of allergen extracts, the molecular weights of a mite and a birch pollen allergen extract were determined using a Bio-Sil TSK 250 column with a guard column. The allergenic activities were measured by RAST inhibition and the antigen/allergen patterns were determined by crossed (radio)immunoelectrophoresis. The original extracts and the corresponding eluates after HPLC runs showed the same allergenic activity and the same number of antigen/allergen precipitation lines. Only slight differences in the peak heights of some precipitates were observed.

Allergens↗
PubMed1986

Kinetics of allergen release from house dust mite Dermatophagoides pteronyssinus.

Purified bodies of Dermatophagoides pteronyssinus were extracted over different extraction times. The release kinetics of proteins, carbohydrates, and allergens were assessed by different methods. Additional attempts have been made to elucidate the extraction process also on a molecular basis. For this purpose the extracts were analyzed by isoelectric focusing, SDS gel electrophoresis, crossed radioimmunoelectrophoresis, and an immunoprint technique. Carbohydrates, proteins, and allergens were released immediately from the mite bodies but demonstrated different release kinetics. The release of carbohydrates and proteins was linear up to 48 hours but demonstrated different slopes. The major allergen P1 demonstrated steady release up to 48 hours, whereas the overall allergenic activity as determined by RAST inhibition demonstrated typical saturation kinetics reaching a plateau after 4 to 5 hours of extraction. It was demonstrated by parallel-line assay of the RAST-inhibition curves that the allergenic determinants remained unchanged up to 48 hours. Different electrophoretic methods demonstrated that all relevant proteins and allergens were released immediately from the mite bodies. The electrophoretic patterns changed only slightly up to 4 hours. Prolonged extraction times up to 48 hours did not affect the patterns. This makes a loss of proteins or allergens by degradation unlikely.

Allergens↗
PubMed1986

Contact urticaria to rubber gloves is IgE-mediated.

Serological studies were performed in 3 patients with typical contact urticaria to latex surgical gloves. Specific IgE antibodies to natural latex as well as to vulcanized glove material have been detected by radioimmunoassay. Contact urticaria to latex is considered to be of the immunologic type.

Adult↗
PubMed1986

Quality of timothy pollen (Phleum pratense) from different pollen seasons and different suppliers.

We investigated seven batches of timothy (Phleum pratense) pollen from four different pollen seasons and three different manufacturers in the USA and Europe by several biochemical and immunochemical methods. By measuring the contents of hexoses, proteins and the total allergenic activity using RAST inhibition we could not find any significant differences in quality between pollen of different seasons, different geographical origin and different manufacturers. There were only slight differences in the staining of some protein bands and in peak height of some antigens in isoelectric focusing (IEF), SDS-PAGE and crossed immunoelectrophoresis. These differences in IEF and SDS-PAGE patterns seem to be specific for the pollen samples from the USA and Europe, respectively. We also observed minor differences in IEF immunoprint patterns, mainly for some basic proteins. The allergen patterns in crossed radioimmunoelectrophoresis were very similar, with only minor differences in concentration of the basic allergens. It appears that timothy pollen from different years can be produced in different geographical areas by different manufacturers with fairly constant allergenic activity.

Carbohydrates↗
PubMed1985

Ultrasound diagnosis of C-cell carcinoma of the thyroid.

Ultrasound screening in 9 and 21 members, respectively, of two families affected by familial C-cell carcinoma, as well as sonographic findings in 6 patients with sporadic medullary carcinoma of the thyroid (MCT), are reported. Unilateral and bilateral tumor nodules were identified by sonography in 12 of a total of 13 patients with MCT (n = 9) or local tumor recurrence following thyroidectomy (n = 4); one carcinoma, 3 X 6 mm in diameter, was missed. Nine of the 12 (75%) were clinically occult, nonpalpable C-cell carcinomas. The smallest occult MCT correctly diagnosed by ultrasound had a diameter of 4 mm. A positive correlation was found between the sonographically determined tumor mass (number/size of nodules/metastases) and the plasma calcitonin level. Pheochromocytomas were diagnosed by ultrasound as part of a multiple endocrine neoplasia (MEN IIa) in two patients with MCT. The echomorphologic findings of the intrathyroid C-cell carcinomas and their metastases display characteristic but nonspecific features. Sonographic findings on these tumors should therefore not be interpreted without consideration of plasma calcitonin assays.

Adolescent↗
PubMed1985

[High resolution real time sonography of parathyroid tumors].

A total of 15 patients with parathyroid tumours were scanned preoperatively using high-resolution real-time sonography (7 MHz). Of these, 14 had a single adenoma or primary hyperplasia, one had a carcinoma. In 13 out of 15 patients the echogenicity of the parathyroid tumours was consistently less than of the adjacent normal thyroid tissue; in 2 out of 15 cases parathyroid neoplasms were barely distinguishable in echogenicity from thyroid tissue. No specific echomorphologic differences were proved to exist between primary parathyroid hyperplasia, adenoma or carcinoma. The smallest parathyroid neoplasia in this study detected by sonography measured 5 x 8 mm in diameter. The value and limits of parathyroid sonography were discussed with regard to our own experience and the reports in the literature.

Adenoma↗
PubMed1985

Comparison of classical Lowry, modified Lowry and a dye-binding assay for the estimation of protein in allergen extracts and influence of different parameters on the modified Lowry assay.

Protein values of dialysed allergen extracts determined by Lowry, modified Lowry (trichloroacetic acid precipitation of the proteins) and dye-binding assay were compared. The influence of different parameters on the modified Lowry was examined. The reproducibility of the modified Lowry was checked with three independent measurements. For the examination of recovery a constant amount of 6-grass pollen allergen proteins was added to the samples of the standardized human serum albumin prepared for the calibration curve. The samples were measured by modified Lowry. The mean of the ratio between the protein values of the dialysed allergen extracts obtained by modified Lowry and those obtained by classical Lowry was 3.59 (coefficient of variation Cv = 45%). The mean of the ratio between the protein values of the allergen extracts obtained by modified Lowry and dye-binding assay was 1:0.71 (Cv = 31%). Phenol interfered with the modified Lowry. Phenolic allergen extracts showed higher "protein values" than non-phenolic allergen extracts. This influence could be reduced by a second precipitation of the dissolved precipitate. The precipitation of non-phenolic dialysed aqueous allergen extracts was complete after the first trichloroacetic acid precipitation. By incubating samples with the Folin-Ciocalteu's reagent at 55 degrees C in a waterbath, the time necessary for developing the colour could be reduced from 45 min to 5 min. Protein measurements by modified Lowry of a 6-grass pollen allergen extract in three different laboratories showed good reproducibility. For these extract 785 micrograms protein/ml (Cv = 4%) could be measured.(ABSTRACT TRUNCATED AT 250 WORDS)

Allergens↗