Single Precipitate Research Articles

Comparative Studies on Tree-Pollen Allergens

The structural changes as due to thermal denaturation of antigens and allergens of birch pollen produced by gel filtration chromatography were studied by circular dichroism, RAST inhibition and immunoelectrophoretic techniques. Results of circular dichroism showed that both BV3 and BV4 contained 20–25% alfa-helix, while BV45 consists of beta-pleated sheet and random coil. Fractions BV3 and BV4 lost about 50% of their native tertiary structures when heated at 100<sup>o</sup>C for 3 h. The number of antigenic lines found in the CIE plates for the heat-treated fractions was reduced as well. The CRIE preparations showed unchanged radio-staining for the heated BV3 fraction (correlated to the untreated BV3 fraction). For the BV4 fraction a diminished radiostaining was observed. A reduction of 8-fold and 4-fold of the IgE binding capacity was observed in RAST inhibition for fractions BV3 and BV4, respectively after heating. The purified fraction BV45 showed 50% decrease in CD absorption after heating to 100°C for 1 h. The CIE pattern of the heated fraction gave a single precipitate line and the area under the precipitate line was reduced as compared to the unheated fraction. The CRIE plate showed reduced intensity of radiostaining. Skin prick tests and PK tests confirmed the reduced allergenicity of the heated fractions. These findings confirm the relative thermostability of the allergens in birch pollen. However, heating infers irreversible changes on the tertiary structure of the molecules, and these structural changes can be correlated to a quantitative reduction of allergenic reactivity.

Initiation and growth of the grain boundary discontinuous precipitation reaction

Evidence is presented indicating that within the Cu-Be alloy system, which undergoes a discontinuous precipitation reaction, the aging temperature can affect the initiation mechanism of the reaction. At low temperatures (below ~ T m /2) precipitates induce grain boundary migration by a puckertype process. At high temperatures (above ~ T m 2) thermal migration of grain boundaries precedes boundary precipitation. These observations are used to support a proposed explanation for the occurrence of both single seam and double seam precipitate morphologies in the growth stages of the reaction in aged alloys. The double seam morphology which is the favored mode of growth at low temperatures is shown to arise as a result of the ‘S’ mechanism previously proposed as a mechanism for discontinuous coarsening. The single seam morphology which predominates at high temperatures is due to boundary migration in one direction only.

The immunochemical identification of a thiol-protein disulfide oxidoreductase (glutathione-insulin transhydrogenase) in pancreatic islets

Pancreatic islets contained insulin-degrading activity that was completely removed by antisera to purified microsomal thiol-protein disulfide oxidoreductase from rat liver. In Ouchterlony double-diffusion experiments with these antisera, extracts of islet homogenates showed a single precipitation band of identity with the purified liver enzyme. Twodimensional immunoelectrophoresis also gave a single precipitate peak like that of the liver enzyme. The concentration of the enzyme in rat islets as determined by quantitation of the precipitates obtained in the electroimmunodiffusion analysis was in the order of 1.0% of total islet protein. The results suggest that, in vitro, cleavage of insulin into its polypeptide chains is catalyzed by the thiol-protein disulfide oxidoreductase. This enzyme promoting thiol-protein disulflde interchange may be important for regulating the content of pancreatic insulin.

The bile-salt-stimulated lipase in human milk. Purification and characterization.

The bile-salt-stimulated lipase was purified from human whey by chromatography on heparin-Sepharose and Affi-Gel blue. The purified enzyme gave a single band with a molecular weight of 90 000 on dodecylsulphate/polyacrylamide gels and this band accounted for at least 98% of the protein on the gel. An antiserum to the purified lipase completely inhibited the enzyme activity and gave a single precipitate against human whey and purified lipase. The bile-salt-stimulated lipase was inhibited by diisopropylfluorophosphate, which bound to the purified enzyme in a molar ratio of 0.85 mol/mol. The lipase is a glycoprotein with a high content of acidic amino acid residues and an isoelectric point of around 4. Proline constitutes more than 10% of the total amino acid residues. The purified lipase has a turnover number of around 150 S-1.

The use of specific antiserum induced by lectin-antigen complexes to investigate the outer membrane antigens of Neisseria gonorrhoeae

Extracts of gonococcal surface antigens, examined by crossed affinity electrophoresis (CAE) with wheat germ agglutinin (WGA), yielded a single antigen-lectin precipitate in agarose gels. This precipitate induced in rabbits a potent antiserum specific for the gonococcal antigen which reacted with WGA. Immune electron microscopy on whole gonococci showed that this antiserum reacted with outer membrane vesicles. Lipopolysaccharide (LPS) purified by phenol-water extraction of whole gonococci reacted with WGA and also with the antiserum to the WGA-antigen complex. This indicated that the antiserum was specific for antigen(s) of the outer membrane complex.

Purification and characterization of Aleutian disease virus.

Virus was isolated from infected mink organs by a combination of tissue homogenization, fluorocarbon extraction and ultracentrifugation. The final preparation was analysed by crossed immunoelectrophoresis and electronmicroscopy. Virions had a capsid diameter of 22 nm. Preparative agarose electrophoresis separated virions from contaminating ferritin. Crossed immunoelectrophoresis of virus gave a single precipitate with sera from infected mink. Crossed immunoelectrophoretic analysis with intermediate gels showed that a part of the virus preparation was complexed with antibody. Serum from a certain mink was found to contain precipitating antibody to (poly)nucleotid. Virus and virus-antibody complexes were found to focus at pH 4.0-4.4 in isoelectric focusing. In SDS-polyacrylamide gel-electrophoresis the main virus protein was found to have a molecular weight of 69000. This study gives further support to the classification of aleutian disease virus as a parvovirus.

Isolation of Late Complement Components by Affinity Chromatography: I. Purification of the Human Complement Component C9 and Production of a C9-Defective Human Serum

A new procedure for the isolation of the human complement component C9 is described. This procedure offers the possibility to prepare functionally pure C9 in a one-step procedure with a high recovery of 10–22% of the biological activity. The 125iodinated C9 had a molecular weight of 78,000 daltons and was only contaminated in traces with other proteins. Further purification by absorption with an «anti-impurity» column lead to a C9 preparation which behaved as a homogenous single polypeptide chain in SDS polyacrylamide gel electrophoresis after reduction with mercaptoethanol. It formed a single bell-shaped precipitate in crossed immunoelectrophoresis with antibodies against human serum in the gel of the second dimension. The recovery of the biological activity after the second purification step was in the order of 6–10%. Both preparative steps could be performed within a few hours. 450 μg C9 protein were isolated from 135 ml human serum. A human serum completely defective in C9 was prepared by the extensive absorption of a smaller volume of human serum proteins with the anti-C9 column.

TEM - characterization of precipitates in Al-Zn5-Mgl

Various techniques of TEM are described to investigate precipitates in metals, which reach dimensions from a few nm to more than 100 nm. The examples presented here are characterized by strong interactions of the lattice structures of the inclusions and the matrix, therefore replica techniques are not applicable (an example of that is presented in [1]] because the metastable phases might change.Selected area diffraction of regions of about 100 nm give well defined diffraction patterns of the phases present in that part of the specimen. Generally the diagram corresponding to a single precipitate cannot be obtained, but in contrast to microdiffraction techniques sharp diffraction spots can be measured. The orientation relationship between the inclusions and the matrix can be studied best, if the beam direction coincides with the main cristallographic zone axis, i.e. in cubic lattices: <001>, <112>, <111>, <011>. At these poles the patterns are highly symmetric, especially because of doublediffraction, but the diagram can be constructed to unfold the coinciding patterns.

OSTWALD RIPENING IN IONIC CRYSTALS

OSTWALD RIPENING IN IONIC CRYSTALS

Martensitic Transformation of Iron Precipitates in a Copper-2 wt.-% Iron Alloy

Transformation characteristics of iron precipitates formed in a Cu-2 wt.-%Fe alloy aged at 800°C have been investigated by means of high-voltage electron microscopy, especially by utilizing the selected micro area electron-diffraction technique which makes it possible to take diffraction patterns from within a single precipitate. In the iron precipitates a banded structure is observed with alternate light and dark contrasts, and is unambiguously verified to consist of alternate lamellae of two bee martensite regions which are twin-related to each other with respect to a (112) plane. It is thus clearly shown that the iron precipitates completely undergo martensitic transformation, even when they are embedded in an inert matrix. Thinning of specimenscan also promote martensitic transformation as well as deformation and the extraction of precipitates from specimens. The orientation relationship between the martensite (α′) and surrounding matrix (E) crystals is determined to be (111)ɛ|(011)α′, [101]ɛ‖[111]α′; this is the Kurdjumov-Sachs orientation relationship, inagreement with previous work by Easterling and Weatherly.

Cellular immune activity of a galactomannan-protein complex from mycelia of Histoplasma capsulatum.

A galactomannan-protein complex was extracted from defatted mycelium of Histoplasma capsulatum with 0.25 N alkali at 25 C. It accounted for 7.5% of the cell dry weight in nondialyzable form and had a galactose-mannose ratio of 2:5. The complex containing 68% protein was separated into polysaccharide and glycoprotein components by ion-exchange chromatography. The galactomannan formed a single precipitate in immunodiffusion, sharing a common antigen with the glycoprotein which contained two precipitating antigens. The glycoprotein elicited delayed-type hypersensitivity in guinea pigs as measured by skin-test and macrophage migration inhibitory factor assay, with 5 mug resulting in 86% specific inhibition. The galactomannan was skin-test negative, but 10 mug resulted in 80% specific inhibition of migration.

Cross-reactivity of synthetic linear dextran with anti-B512 dextran. Viewpoints on the nature of the antigenic determinants of dextran.

A fraction of synthetic linear dextran (D-8) of Mn 26,900 was tested for cross-reactivity with rabbit anti-B512 dextran antiserum (RAD) and compared to a B512 dextran fraction (D-40) of similar molecular weight distribution. B512 dextran is a linear α-1–6 linked polyglucose with few side chains: about one branch per 20 glucose units; at least 80% of branches comprise only 1–2 glucose residues. The synthetic dextran is a completely linear α-1–6 linked polyglucose. In Ouchterlony gel double-diffusion, both D-8 and D-40 produced single precipitate lines; in a comparative array, complete fusion of precipitate lines of D-8 and D-40 indicated the identity of antigenic determinants. In quantitative precipitation and single radial immunodiffusion, D-8 and D-40 precipitated comparable amounts of antidextrans and produced precipitate haloes of similar size. Systemic lethal anaphylaxis could be elicited by D-8 in passively sensitized guinea pigs, and protection from lethal shock was obtained by adding isomaltooctaose to D-8 upon challenge. Similarly, passive cutaneous anaphylaxis (PCA) of comparable intensity was elicited by D-8 and D-40 upon challenge of sensitized guinea pigs. Neutralization of PCA by adding D-8 to RAD prior to intradermal sensitization and challenge with D-40 also indicated the identity of antigenic determinants. These findings are taken as evidence that D-8 is a homodeterminant antigen with many determinants, and that the antigenic determinants of both linear synthetic and B512 dextran are repetitively arranged along the molecular chain.

Behaviour of Leal Ion Sensitive Chalcogenide Electrodes in Buffered Lead-Ion Solutions

Abstract The response of single crystal and precipitate lead sensitive chalcogenide electrodes (PbS, PbSe, PbTe) in lead ion buffer solutions was studied. It was found that the sensitivity toward Pb decreased in the order PbS∼PbSe > PbTe, while the pH sensitivity follows the reverse order: PbS<PbSe» PbTe. Ag2S present in the electrode affects the behavior of the electrode and causes a sharp decrease in the electrode sensitivity when present in a concentration greater than 90% wt. The chalcogenide electrodes are not very advantageous as direct potentiometric sensors but they can be used advantageously in precipitation and complexometric titrations of lead.

Antigenic profiles of proteinpolysaccharide complex from human costal cartilage. I. Antigenicity of sialoprotein moiety.

Antisera, obtained by immunization of rabbits with proteinpolysaccharide complex of human costal cartilage, produced at least two zones of precipitate with antigen on the Ouchterlony assay. The major proteinpolysac-charide fraction, obtained from a DEAE cellulose column chromatography and from a gel filtration, produced a single precipitate line which fused with that of bovine material and therefore was nonspecific between human and bovine material. On the electrophoresis in an agarose gel as well as in a Pevikon powder block, a discrepancy in a mobility occurred between the major glycosaminoglucuronoglycan-containing spot and the antigenic moiety. This antigenic material possessed the chemical composition characteristic to the glycosaminoglucuronoglycan-core protein complex as well as the sialoprotein. After hyaluronidase digestion, this antigenic fragment was characterized rather as a sialoprotein.

The interaction in austenitic stainless steel between the stabilizing elements and carbon or nitrogen and its effect on corrosion behaviour

Though “weld decay” in austenitic CrNi steels can be eliminated by the use of stabilizing elements to lock up the carbon and nitrogen, related effects are still observed under onerous conditions, e.g. exposure to hot nitric acid. A simple model is developed in which it is assumed that the behaviour of an alloy depends on the concentrations in solutions of two elements which form a precipitate of form MX and the amount of precipitate formed, and that the concentrations in solution are given by the dissociation constant of the precipitate. The model is extended to cover the cases where a single precipitate of form M iX j occurs, and where a third element Y also reacts with M so that two precipitates MX and MY are formed. Existing data on the corrosion behaviour of austenitic CrNi steels are considered in terms of the model. The model provides a satisfactory fit of the data and explains many of the known facts about the behaviour of these alloys after various heat treatments and the relative performance of titanium-and niobium-stabilized steels.