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J Steensgaard

Publications and source records attributed to J Steensgaard.

At least 19 recordsLinked to original sources
PubMed1992

GTP-binding proteins in luminal and basolateral membranes from pars convoluta and pars recta of rabbit kidney proximal tubule.

The GTP-binding proteins on luminal and basolateral membrane vesicles from outer cortex (pars convoluta) and outer medulla (pars recta) of rabbit proximal tubule have been examined. The membrane vesicles were highly purified, as ascertained by electron microscopy, by measurements of marker enzymes, and by investigating segmental-specific transport systems. The [35S]GTP gamma S binding to vesicles, and to sodium cholate-extracted proteins from vesicles, indicated that the total content of GTP-binding proteins were equally distributed on pars convoluta, pars recta luminal and basolateral membranes. The membranes were ADP-ribosylated with [32P]NAD+ in the presence of pertussis toxin and cholera toxin. Gel electrophoresis revealed, for all preparations, the presence of cholera toxin [32P]ADP-ribosylated 42 and 45 kDa G alpha s proteins, and pertussis toxin [32P]ADP-ribosylated 41 kDa G alpha i1, 40 kDa G alpha i2 and 41 kDa G alpha i3 proteins. The 2D electrophoresis indicated that Go's were not present in luminal nor in basolateral membranes of pars convoluta or pars recta of rabbit proximal tubule.

Animals↗
PubMed1991

Hydrocolloid occlusion for the treatment of neurovascular corns.

The goal of this study was to determine the effect of hydrocolloid occlusion on neurovascular corns. The design was an observer-blinded, randomized, controlled study. Thirty consecutive patients participated in the trial. The patients received curettage alone or curettage with hydrocolloid occlusion. Six treatments were given over 12 weeks. A follow-up examination was performed 3 months after termination of the trial. Outcome measures were the size of the corns, a discomfort score, and an overall judgment of the trial. The results demonstrated no benefit of occlusion for symptoms or signs of neurovascular corns. The patients treated with occlusion were, however, generally more satisfied than the conventional group.

Aged↗
PubMed1984

Binding properties of monoclonal anti-IgG antibodies: analysis of binding curves in monoclonal antibody systems.

The binding properties of an immune complex-forming system comprising human IgG and mouse monoclonal antibodies against human IgG have been studied. A refined binding assay has been applied directly on ascitic fluid containing monoclonal antibody. Complete sets of binding data of a series of different monoclonal antibodies were collected and analysed by various graphical and statistical methods. Special attention was given to methods which allow determination of specific monoclonal antibody concentration as well as antibody affinity. It was found that the formation of genuine antigen: antibody complexes per se gives rise to deviations from expected linearity in commonly used binding equations. Good correlation was found between the antibody concentrations obtained by various graphical approaches, whereas the size of the association constant seemed to depend on the method in use. The binding pattern was found to be dependent on the concentration of antibody. Most reliable parameters were obtained if the product of the antibody concentration and the association constant was below 10.

Animals↗
PubMed1983

Molecular distribution of preformed thyroglobulin immune complexes.

Thyroglobulin anti-thyroglobulin immune complexes with human antibodies from four patient sera were performed in a wide range of antigen: antibody ratios and run in 5%-40% sucrose gradients. The fractionation showed simple distributions of thyroglobulin molecules in immune complexes, especially in far antigen and far antibody excess. A computer simulation analysis which utilized the experimental presumptions and assumed an association constant in the order of 10(9), showed a large degree of similarity to the experimental results. Thus, a thermodynamically simple explanation of immune complex formation did not contradict the experimental thyroglobulin anti-thyroglobulin complex formation.

Antigen-Antibody Complex↗
PubMed1982

Determination of affinity of monoclonal antibodies against human IgG.

A method for determining the affinity and the concentration of mouse monoclonal antibodies against human IgG has been developed. The method comprises two steps. First, monoclonal antibodies are allowed to combine with radioiodinated human IgG. Secondly, mouse monoclonal IgG with and without complexed IgG is precipitated with rabbit IgG against mouse IgG. As the antigen is divalent complexes of varying composition are formed in this system leading to deviations from linearity in plots obtained in commonly employed analytical systems. The theoretical background of these systems has been studied by computer stimulation, and a concentration effect on the formation of immune complexes were demonstrated. The affinities (in terms of the association constants (of 6 monoclonal antibodies were estimated and found to be in the range from 2 X 10(6) M-1 to 5 X 10(8) M-1.

Animals↗
PubMed1982

Theoretical and ultracentrifugal analysis of immune complex formation between monoclonal antibodies and human IgG.

Immune complex formation of four different mouse monoclonal antibodies against human IgG has been studied using analytical zonal centrifugation. A theoretical model has been used to depict thermodynamic ideal immune complex formation of monoclonal antibodies. It was found that the four monoclonal antibodies differed very much with respect to immune complex formation with human IgG. One of the monoclonal antibodies formed immune complexes in agreement with the theoretical model. Another was strongly related thereto. A third monoclonal antibody formed only a single complex and might exhibit a positive co-operativity between its two sites. A fourth formed as excess of a possibly cyclic complex. Thus monoclonal antibodies differ substantially with respect to physical properties adding a new aspect to the problems of antibody heterogeneity. It is moreover found that analytical zonal centrifugation can be used to estimate the number of antigenic determinants and antibody-binding sites thereby assuring whether or not a particular IgG monoclonal antibody has two binding sites.

Antibodies, Monoclonal↗
PubMed1980

Biochemical aspects of immune complex formation and immune complex diseases.

Formation of immune complexes is a normal part of the immune defence against soluble antigens. Immune complexes may nevertheless play a pathogenic role of their own. The present review discusses antigen antibody interactions with special regard to immune complex formation. A new classification of antigen antibody interactions is proposed, based on the antigenic valence. Oligovalent antigens and bivalent antibodies form genuine immune complexes. Experimental observations and theoretical considerations indicate that although a vast variety of complexes is possible genuine immune complex formation is a self-limiting process, so it is typical that the smallest possible complexes are formed in the largest amounts. Formation of immune complexes differs in mechanism from most other biological ligand binding reactions, and the extent to which immune complex formation follows its own laws is discussed. To explain the mechanism of precipitin reactions, a two-stage model is proposed. The outcome of antigen antibody interactions is further complicated because antibody preparations are typically heterogeneous, and the impact of antibody heterogeneity is also discussed.

Animals↗
PubMed1980

Solubility properties of IgG immune complexes. Comparison between the effects of low molecular weight solvents and polyethylene glycols.

A systematic study of the influence of organic solvents on the interaction between human IgG and rabbit anti-human IgG IgG++ has been conducted, using difference turbidity and sucrose gradient centrifugation methods. It was found that very small alcohols like methanol and ethanol increased the turbidity signal, whereas larger alcohols (propanol, isopropanol, propanediol and ethylene glycol) had no influence on the signal. Sucrose, dioxane, dimethylformamide and dimethylsulphoxide decreased the difference turbidity signal. Sucrose gradient centrifugation did not show any influence. Polyethylene glycols were investigated with regard to molecular weights as well as concentrations. In contrast to the low molecular weight solvents, polyethylene glycols with molecular weights from 600 up to 20,000 gave a clearly expressed increase in the development of turbidity as well as in the size of the final turbidity signal. By sucrose gradient centrifugation, it was found that polyethylene glycols of molecular weights from 1000 to 10,000 had a profound effect on the solubility of immune complexes. The influence depended on the molecular weight as well as on the concentration. By use of [14C]-labelled polyethylene glycol it was found that polyethylene glycols did not form stable complexes with immune complexes. Furthermore, it was found that the zoning phenomenon was retained in the presence of small concentrations of polyethylene glycols, but higher concentrations of polyethylene glycols caused even antigen-excess complexes to show turbidity.

Alcohols↗
PubMed1979

A new gradient former and a simplified procedure for zonal centrifugation of immune complexes.

A new gradient former for production of optimized sucrose gradients for B-XIV zonal rotors is described. Optimized gradients are gradients with an initial shelf to provide capacity and a shape that ensures isokinetic sedimentation conditions. By exhaustive computer calculations it has been found that optimized gradients can be produced by a very simple two-compartment gradient former. The design is given. Moreover, graphs for choice of gradient strength and run duration for preparative and analytical runs of samples containing soluble immune complexes are calculated. The new gradient former allows the use of a very simple procedure for zonal centrifugation.

Antigen-Antibody Complex↗
PubMed1979

On the composition of IgG anti-IgG immune complexes.

The formation of different immune complexes of IgG was followed in a system comprising human IgG as antigen and rabbit IgG directed against the Fc portion of human IgG as antibody. Soluble IgG complexes were analyzed by analytical zonal centrifugation. In antigen excess, 16S complexes predominated. 16S complexes are oligomers of IgG, mainly trimers and tetramers. In decreasing antigen excess larger and larger complexes were formed. It was, however, found consistently that oligomers were always formed in the largest amounts. The largest complexes detectable by this method consisted of about twenty IgG molecules. The solubility of different complexes in polyethylene glycol was also studied. Low concentrations of polyethylene glycol preferentially precipitate large complexes. Four and six per cent polyethylene glycol precipitated all types of IgG complexes although not completely. Polyethylene glycol was seemingly not bound directly to soluble immune complexes, but caued otherwise soluble complexes to precipitate by an indirect mechanism.

Animals↗
PubMed1979

A theoretical approach to precipitin reactions: insight from computer simulation.

The theoretical consequences of different hypotheses of the mechanism of precipitin reactions have been evaluated by means of computer simulation. It has been found that the formation of compositionally different complexes in different antigen/antibody mixtures provides a valid explanation of the zoning phenomenon, but this concept fails to explain the absence of free antigen and of antigen in soluble complexes at the point of maximum percipitation. It is found that the following hypothesis provides an improved qualitative and quantitative explanation of percipitin reactions. In the first stage of the total reaction a series of compositionally different complexes is formed. As the second stage of the total reaction two kinds of processes are proposed. Inherently insoluble complexes precipitate causing the remaining soluble complexes to participate in mutual rearrangements to re-establish a new state of equilibrium in the supernatant. The inherently insoluble complexes, moreover, create a hydrophobic phase, distinct from the supernatant and cause the remaining otherwise soluble complexes to distribute themselves between the two phases according to a partition coefficient. A mathematical apparatus to study the consequences of this hypothesis is presented, and it is demonstrated that the features of precipitin curves can be explained nearly completely this way.

Antigen-Antibody Complex↗
PubMed1979

Measurements of precipitin reactions by difference turbidimetry: a new method.

A new method for the measurement of precipitin and flocculation reactions between antibodies and antigens has been developed. The technique, called difference turbidimetry, involves the use of tandem cuvettes providing the opportunity of using separated and unmixed antigen and antibody as blank solutions for spectrophotometric readings in the ultraviolet wavelength range. By use of this technique genuine difference turbidity spectra have been recorded for the reaction between human serum albumin and rabbit-anti-human serum albumin IgG. It was found that difference turbidimetry at low wavelengths (e.g. 280 nm) allows the construction of precipitin curves with a very clearly expressed zoning phenomenon at a sensitivity which in terms of antigen and antibody concentrations is more than twice the sensitivity of conventional procedures. It is of special interest that the zone of equivalence differs when the same reaction between an antigen and its antibody is measured by difference turbidimetry, by absorbance of washed and redissolved precipitate, and by amount of precipitated antigen.

Antigen-Antibody Reactions↗
PubMed1979

Fc-mediated immune precipitation. II. Analysis of precipitating immune complexes by rate-zonal ultracentrifugation.

The formation of immune complexes was studied by analytical rate-zonal ultracentrifugation using isomolar solutions of rabbit anti-human serum albumin IgG and of the corresponding F(ab')2 fragments. The F(ab')2 fragments retained full ability to react with the antigenic determinants and to form genuine antigen-antibody complexes. Thus, the difference between the IgG and the F(ab')2 systems was supposed solely to reflect the lack of the Fc portion. The complexes formed with F(ab')2 fragments in the zone of low and moderate antigen excess were found to be distinctively mor soluble than those formed with intact IgG. The data indicated that there were two kinds of precipitating immune complexes, namely antibody-rich and antigen-rich complexes. In the antibody-excess zone and the first part of the equivalence zone the immune complexes precipitated due to their antibody richness. Antigen-rich complexes formed in the zone of low antigen excess precipitate only in the presence of antibody-rich insoluble complexes. It is believed that this type of precipitation was due to an Fc-Fc interaction. This new function of the Fc portion of IgG was designated Fc-mediated immune precipitation.

Animals↗