PubMed Health⌕ Search

Biomedical subjects

C J Strasburger

Publications and source records attributed to C J Strasburger.

105 records · Page 6Linked to original sources
PubMed1992

'Normal' cigarette smoking increases free cortisol in habitual smokers.

In habitual smokers salivary cortisol responses to cigarette smoking were investigated. In the first study, 31 adults assigned to two experimental groups smoked either one or two cigarettes of their preferred brand. Mean salivary cortisol levels were significantly elevated after smoking of two cigarettes. In the second study, 10 smokers and 10 nonsmokers provided saliva samples at 20 min intervals over a 12-hr period. While environmental stimuli were paralleled in both groups overall cortisol output was significantly elevated in the smokers. These data suggest that 'normal' cigarette smoking can increase free cortisol levels.

Adult↗
PubMed1992

Synthesis of a cortisol-biotin conjugate and evaluation as a tracer in an immunoassay for salivary cortisol measurement.

Cortisol 3-(o-carboxymethyl)oxime (C3-CMO) and a commercially available biotin-hydrazide derivative were used to synthesize a C3-CMO-biotin conjugate. C3-CMO was converted into a N-hydroxysuccinimide ester derivative which in a second reaction step was allowed to interact with the hydrazide derivative of biotin. This simple-to-perform synthesis yielded a conjugate suitable for use as a tracer in immunoassays for cortisol measurement. Employing biotin as the primary probe in a competitive solid phase immunoassay allows for variable end point determination by means of commercially available labeled avidin or streptavidin derivatives. Streptavidin-Europium was used in conjunction with the DELFIA-system for time-resolved fluorometric end point measurement (TR-FIA) throughout the study. In addition, colorimetric end point determination (ELISA) using streptavidin-alkaline phosphatase as a secondary probe was established and evaluated. Both forms of this non-isotopic assay showed excellent correlation with a commercially available radioimmunoassay adapted for salivary cortisol measurement. The lower detection limit was 0.43 nM for a 50 microliters salivary sample. The intra-assay coefficient of variation was 6.7, 4.7 and 4.0% at cortisol concentrations of 2.2, 5.5 and 13.2 nM, respectively (n = 37), and the corresponding inter-assay coefficients of variation were 9.0, 8.6 and 7.1% (n = 50). The competitive immunoassay requires 1.5 h incubation time and shows robust and reproducible performance. The C3-CMO-biotin conjugate allows for sensitive and flexible end point determination of salivary cortisol levels in immunoassays.

Biotin↗
PubMed1990

Antigenic epitope mapping of the human growth hormone molecule: a strategy to standardize growth hormone immunoassays.

The antigenic surface of the human growth hormone (hGH) molecule has been characterized by means of monoclonal antibodies (MAbs) and their specific binding sites. From a panel of MAbs raised against the 22 kDa variant of hGH, ten distinct antigenic entities could be identified as binding sites for specific MAbs in an epitope mapping experiment. The experiment was based on combinations of the MAbs in two-site assays. By investigating the cross-reactivity of the MAbs with growth hormone (GH) variants and fragments of hGH, the underlying protein structure of some of the partially overlapping epitopes could be identified. By using antibodies of known epitope specificity as competitors in receptor binding assays of hGH, it was demonstrated that the lactogenic and somatogenic biological effects of hGH are mediated through different portions of the hormone. These observations may explain some of the differences between the various immunoassays for hGH. A strategy for selecting antibodies of known epitope specificity and biological relevance for the design of plasma GH immunoassays is outlined. The results of such an immunoassay should closely reflect the biological activity of circulating plasma hGH.

Animals↗
PubMed1989

Chemiluminescent labelled streptavidin (STAV) as a universal marker in steroid and peptide immunoassays.

The tetrameric structure of streptavidin and its exceptionally strong affinity to biotin (Ka = 10(15)M 1) can be exploited to achieve an amplification of the signal in immunoassays. In the approach described here streptavidin (STAV) labelled with aminobutylethyl-isoluminol (ABEI) served as a universal marker in immunoassays for both haptens and big antigens. The advantageous features of streptavidin can be applied to any immunoassay using biotinylated antibodies as the primary probe. In two-site immunometric assays for larger antigens the liquid phase 'tracer' antibody is biotinylated. In hapten assays the solid phase antigen technique (Wood et al., 1982) is employed, in which sample-antigen and solid phase-antigen compete for a biotinylated antibody. In this paper we demonstrate the use of STAV-ABEI as a universal chemiluminescent label in steroid assays and in an immunometric assay using human growth hormone (hGH) as an example.

Antigens↗
PubMed1989

Cortisol and behavior: 1. Adaptation of a radioimmunoassay kit for reliable and inexpensive salivary cortisol determination.

We adapted a commercial serum cortisol radioimmunoassay (RIA) kit for use with saliva specimen. Using 50 microliters sample volume, the lower sensitivity was found to be 0.02 microgram/dl with intraassay variation coefficients between 5.4 and 8.9% at different concentrations. The 50% intercept was either 0.5 or 0.26 microgram/dl (50 or 100 microliters standard/sample volume). Fifty-four early morning samples from healthy adults showed absolute concentrations which are closely comparable to respective data from other laboratories. A comparison of 35 saliva samples which were each assayed with the adapted RIA as well as with three other commercial kits revealed high correlations between these assays (r = .94 to r = .97). Data on salivary cortisol responses to CRH stimulation and dexamethasone suppression in healthy subjects further the validity of the assay results. The most important contribution of this assay modification, however, is thought to be its impact on analysis costs: The protocol presented in this paper allows for reliable salivary cortisol measures with a reduction of costs for analytical material to 25% compared to serum determinations.

Corticotropin-Releasing Hormone↗
PubMed1989

Indication of different lactogen and somatogen binding sites in the human growth hormone molecule as probed with monoclonal antibodies.

The relationship between the structure of human growth hormone (hGH) and the hormone-receptor interaction has been investigated using as probes monoclonal antibodies (Mabs) to hGH of defined epitope specificity profile. Seven high affinity Mabs were studied for their ability (i) to inhibit the binding of 125I-hGH to Nb2-SP rat lymphoma cells and to IM-9 human lymphocytes possessing lactogen and somatogen type receptors, respectively; and (ii) to interfere with the hormone (hGH or Met8Leu hGH)-induced proliferation in Nb2-11C lymphoma cells. The ability of these Mabs to inhibit the 125I-hGH binding and the hormone-induced proliferation in Nb2-11C cells was negatively correlated with the ability of these Mabs to cross-react with met14 hGH. Furthermore, Mabs Nos. 3 and 7, which cross-reacted minimally (0.2-0.4%) with Met8Leu hGH, were unable to interfere with the mitogenic activity of Met8Leu hGH in Nb2-11C cells. These results indicate that the first 13 amino acids of the N-terminal region of hGH are necessary for its lactogen activity. The inhibition of 125I-hGH binding to IM-9 cells by these Mabs was similar to those observed in Nb2-SP cells, except for Mabs Nos. 19 and 1. These Mabs inhibited more strongly the binding of 125I-hGH to IM-9 than to Nb2 cells and recognized antigenic epitopes close to the C-terminal part of the molecule. These results suggest that the somatogen receptor binding site of hGH may be located on two sites, one at the N-terminal and the other one close to the C-terminal, while the lactogen receptor is mainly confined to the N-terminal part.

Animals↗
PubMed1989

The antigenic epitopes of human grown hormone as mapped by monoclonal antibodies.

The antigenic epitopes of human GH (hGH) have been investigated using monoclonal antibodies (Mabs) to hGH. A panel of 12 Mabs was incorporated into two-site binding assays in order to investigate the antigenic surface of hGH. Nine reaction patterns were observed. The Mabs were further characterized in terms of their cross-reactivity with human placental lactogen, human PRL, with recombinant hGH molecules (MetLeu8hGH, Met14hGH), and with fragments derived from enzymatic or chemical digestion of hGH. These studies provided information on the antigenic sites that are shared with human placental lactogen and on the N-terminal and C-terminal regions of the hGH molecule. Based upon these findings, we tentatively suggest the epitope model for hGH comprising at least 10 antigenic sites (epitopes) which may be grouped into five antigenic regions.

Animals↗
PubMed1985

Ceruloplasmin and transferrin in human seminal plasma: are they an index of seminiferous tubular function?

Transferrin and ceruloplasmin have been measured by a solid-phase chemiluminescent method in seminal fluid and circulating blood of normal and vasectomized subjects (1 year after operation). This study has confirmed that approximately 80% of seminal transferrin comes from the testis, while seminal ceruloplasmin was not found different in the two groups. In patients affected by azoospermia due to seminiferous tubular damage (n = 15) in whom an obstruction was previously excluded, seminal transferrin was always below the normal range. On the contrary, seminal ceruloplasmin was always in the normal range, and circulating follicle-stimulating hormone was found above the normal range only in nine cases. No correlation was found between seminal transferrin and circulating follicle-stimulating hormone in such groups. In an unselected group of infertile patients with decreased sperm concentration and/or sperm motility, seminal transferrin was found correlated with the sperm count. These studies seem to suggest that seminal transferrin is a reliable index of seminiferous tubular function.

Ceruloplasmin↗
PubMed1984

Synthesis and evaluation of luminescent tracers and hapten-protein conjugates for use in luminescence immunoassays with immobilised antibodies and antigens. A critical study of macro solid phases for use in immunoassay systems, Part II.

This article describes the synthesis of labels and hapten-protein conjugates for use in bio- and chemiluminescent immunoassay systems, together with the problems encountered. The effects of maleimide upon acetate-, adenylate- and pyruvate kinase activity have been studied, as well as upon the luciferin-luciferase monitoring system. Maleimide inhibited both acetate and adenylate kinase but showed no inhibition of pyruvate kinase and the monitoring reagent. Four heterobifunctional reagents were tested for their capability in forming pyruvate kinase-donkey-anti-rabbit IgG conjugates which retained enzyme and antibody activity. The best results were obtained with succinimidyl-4-(N-maleimidomethyl)-cyclohexane-1-carboxylate and succinimidyl-6-(p-maleimidophenyl)-hexanoate. The relationship between the amounts of succinimidyl-4-(N-maleimidomethyl)-cyclohexane-1-carboxylate and IgG was studied with respect to enzymic activity of the conjugate. The Michaelis-Menten constants for both conjugated and non-conjugated pyruvate kinase were calculated and compared. It was found that the maximal velocity (Vmax) of the conjugated enzyme was lower than that of the non-conjugated enzyme although the "apparent" Km value was the same for both conjugated and non-conjugated pyruvate kinase. The pyruvate kinase-anti rabbit IgG conjugate was tested for its ability to bind to rabbit-IgG coated polystyrene balls. In addition to bioluminescent labels, the synthesis of chemiluminescent markers was undertaken and optimised. The three substances used for labelling were diazoluminol, diazoisoluminol and N-(4-aminobutyl)-N-ethylisoluminol hemisuccinamide the latter being used as an N-hydroxysuccinamide "active" ester. The ratio of label to IgG was studied for diazoluminol and N-(4-aminobutyl)-N-ethylisoluminol hemisuccinamide active ester after it had been discovered that diazoisoluminol was not suitable for coupling to antibodies. The optimal molar ratios label: IgG were for diazoluminol 40:1 and for N-(4-aminobutyl)-N-ethylisoluminol hemisuccinamide active ester 60:1. Increasing the substitution rate led to a lessening of the dynamic range, shown by an increase in the ratio between unspecific binding (noise) to maximal binding (signal) in an assay. The synthesis of hapten-protein conjugates for covalent coupling to polystyrene balls was undertaken as this formed part of the preparation for the assays described in Part III. The optimal production of gentamicin-bovine serum albumin and thyroxine-transferrin conjugates has been described in detail.

Animals↗
PubMed1984

An evaluation of four different luminescence immunoassay systems: CELIA (chemiluminescent immunoassay), SPALT (solid-phase antigen luminescence technique), ILMA (immunoluminometric assay) and ILSA (immunoluminometric labelled second antibody). A critical study of macro solid phases for use in immunoassay systems, Part III.

The performance of different solid-phase luminescence immunoassays has been documented using four different assay concepts. These are CELIA (chemiluminescence immunoassay), SPALT (solid-phase antigen luminescence technique), ILMA (immunoluminometric assay) and ILSA (immunoluminometric labelled second-antibody assay). CELIA is analogous to a solid-phase radioimmunoassay and uses a labelled antigen, SPALT and ILSA use a labelled second (species-specific) antibody and ILMA a labelled substance-specific antibody, i.e. analogous to the immunoradiometric assay. Both bioluminescent and chemiluminescent labels have been used. Pyruvate kinase was used for bioluminescence and diazoluminol and N-(4-amino-butyl)-N-ethyl isoluminol hemisuccinamide for chemiluminescence. Relevant quality-control parameters and reference ranges have been given for the optimised assays. Assays described are: thyroxine, thyroxine binding globulin, cortisol, caeruloplasmin, ferritin and C-reactive protein. Luminescence immunoassays with coefficients of variation comparable with radioimmunoassay have been designed, values of under 5% being obtainable within the working range of the assay.

Animals↗
PubMed1982

Solid phase antigen luminescent immunoassays (SPALT) for the determination of insulin, insulin antibodies and gentamicin levels in human serum.

We describe an interesting and novel alternative to conventional immunoassay techniques for the measurement of antigens and antibodies in body fluids. The label used for all assays is a pyruvate kinase-IgG conjugate of the relevant second (species-specific) antibody. All assays follow the same principle in which a solid phase antigen is used to adsorb unreacted first (substance-specific) antibody following a conventional antibody-antigen reaction in a liquid phase. After washing, the solid phase antigen-first antibody is allowed to react with the labelled second antibody. The solid phase is then washed and the pyruvate kinase bound to the solid phase is used to generate ATP which is measured kinetically in a luminometer. Assays are described for insulin, insulin antibodies and gentamicin to demonstrate both the versatility and sensitivity of this type of assay. The insulin assay had a lower detection limit of under 0.25 microU per tube and was comparable with the radioimmunoassay used for routine purposes both in sensitivity and reproducibility. The insulin antibody assay correlated well with the radiometric determination used routinely in the laboratory. The gentamicin assay correlated well with the routine commercial radioimmunoassay and also had comparable coefficients of variation. In all cases, the inter- and intra-assay variation was under 10% in the range of interest.

Animals↗
PubMed1982

Enzyme enhanced luminescence immunoassay for the determination of transferrin concentrations in serum.

An enzyme enhanced luminescence immunoassay has been developed using the determination of serum transferrin as a model. The assay has been compared with an established radioimmunoassay, the details of which have already been published (Wood, W. G. et al. (1980) Fresenius Z. Anal. Chem. 301, 120). After comparison, the luminescence immunoassay was introduced as the routine method for determining transferrin levels in human serum. The enzyme enhanced luminescence immunoassay uses a pyruvate kinase labelled transferrin as tracer, which, after the antibody-antigen reaction and subsequent bound-free separation, is used to generate adenosine triphosphate (ATP) from adenosine diphosphate(ADP) and phosphoenol pyruvate. The ATP is then monitored with a firefly luciferin-luciferase reagent, using a luminometer as detection system. The intra- and interassay coefficients of variation were under 10% and under 13% respectively in the range 0.6-4.5 g . liter-1. The stability of the lyophilised tracer was in excess of 5 months when stored at 4 degrees C, and the sensitivity of the assay lay under 0.1 micrograms per tube, although the detection limit of this type of assay system has not been fully explored.

Adenosine Triphosphate↗
PubMedDate not supplied

Doping with growth hormone.

Triggered by the Olympic games in Sydney last year, many articles in the press suggested that recombinant human growth hormone (hGH) is one of the most popular performance enhancing drugs used by athletes. However, any hard facts on hGH abuse were provided by Australian customs officers--and not by laboratory assessment. The lack of an official test for hGH doping together with the widespread rumours on its tremendous beneficial effects seem to make this compound attractive for athletes. From a scientific point of view, there are two major questions about this issue: First, as there is no controlled study demonstrating a profound effect of hGH administration on workload capacity in healthy adults, why do athletes use hGH? Second, how could the application of a substance naturally occurring in the human body be detected? Both aspects are discussed in this article.

Australia↗