Effect of milk on surface properties of Staphylococcus aureus from bovine mastitis

Abstract The surface hydrophobicity of cells of Staphylococcus aureus strains isolated from bovine mastitis grown on conventional agar and broth media was drastically reduced after incubation with bovine milk. Strains grown in high carbohydrate-high salt media yielded cells with reduced surface hydrophobicity compared to cells grown in conventional media, and adding bovine milk to minimal medium also yielded cells with reduced surface hydrophobicity, as determined by hydrophobic interaction chromatography and the salt aggregation test. Incubation of strains in milk and growth in a medium supplemented with bovine milk also significantly changed bacterial surface charge as determined by free-zone electrophoresis. Strains with high or with decreased adsorptive and aggregating properties did not produce surface capsule or slime. Heat treatment (60° C or 80° C) of the bacterial suspensions did not significantly change their adsorptive and aggregating properties.     

The gene for insulin-like growth factor-binding protein-1 is localized to human chromosomal region 7p14-p12.

Insulin-like growth factors (IGF) I and II are bound to high-affinity binding proteins in the blood circulation and other body fluids. These IGF-binding proteins are expressed at different concentrations in different tissues and are thought to regulate the activity of IGF I and II. Cloned cDNA for IGF-binding protein-1 (IGFBP1) has been used to verify the location of its gene to human chromosome 7 by Southern blotting to DNA from a human-mouse hybrid cell line. Further, by in situ hybridization the gene was regionally localized to 7p14-p12, and a Mendelian-inherited two-allele BglII restriction enzyme length polymorphism was identified, with the most frequent allele occurring in 53% of the chromosomes.     

Immunoabsorption of membrane-specific antibodies for determination of exposed and hidden proteins in human erythrocyte membranes.

Human erythrocyte membrane proteins solubilized with the non-ionic detergent Berol EMU-043 have been characterized by crossed immunoelectrophoresis with rabbit antibodies raised against the membrane material. Three out of sixteen membrane-specific immunoprecipitates disappeared when the antisera were first absorbed with intact erythrocytes. This finding indicates that three antigens are exposed on the outside of the erythrocyte membrane. One of these antigens showed acetylcholinesterase activity, and another was the major glycoprotein (glycophorin) as shown by crossed-line immunoelectrophoresis. No antigenic determinants of the latter protein were detected within the membrane or on its inner surface. In crossed immunoelectrophoresis with antisera after absorption with washed, non-sealed membranes only one precipitate remained. This precipitate corresponded to albumin. Accordingly, several proteins seem to have antigenic determinants exposed on the inside of the membrane.     

Electrophoresis, crossed immunoelectrophoresis, and isoelectric focusing in agarose gels with reduced electroendosmotic flow

An ion exchange method using QAE-Sephadex for preparation of agarose with a low electroendosmotic flow and reduced adsorption properties is described. The successful use of such agarose for the separation of highly cationic proteins is illustrated.A method for isoelectric focusing of proteins in gels made from a mixture of purified agarose and a water-soluble non-cross-linked acrylamide polymer is described. This method can be combined with immunochemical identification by electrophoresis of the separated components into antibody-containing agarose gels, also containing such a polymer of acrylamide.     

Correlation of genes in the pap gene cluster to expression of globoside-specific adhesin by uropathogenic Escherichia coli

Abstract Expression of globoside-specific pilus adhesin of Escherichia coli is the virulence factor most commonly associated with pyelonephritis. In the clinical isolate J96 (O4:K6:H5) expression of globoside binding pili require the proteins encoded by the papE, papF , and papG genes in the pap gene cluster. Probes derived from these genes were used in dot blot hybridization analysis of E. coli urinary tract isolates obtained from patients with significant bacteriuria. Fecal E. coli isolates from healthy individuals were also analyzed. The probe encompassing the papF and papF J96 genes hybridized to all urinary tract infectious (UTI) isolates expressing globoside-specific adhesin, whereas papG J96 only hybridized to the strain from which the fragment was cloned. In contrast, a papG -specific probe from the O:6 strain IA2 hybridized to all but one of the UTI isolates that expressed the adhesin. In both materials, but especially among the fecal isolates, strains were found that hybridized to the probes but did not express the adhesin. The data shows that papEF -specific DNA can be used for the diagnosis of potentially pyelonephritic E. coli .     

Simplified luciferase assay of NAD+ applied to microsamples from liver,kidney and pancreatic islets

A new single-step procedure for the bioluminescence assay of NAD⁺, permitting measurements on the pmol level (10^?12 mol) is described. Acid extracts of NAD⁺ were prepared in different tissues. The acidification destroys reduced pyridine nucleotides and most enzymes which are present in the tissue sample. After neutralization the extract is added to a light-yielding solution, and the luminescence is measured with a photomultiplier. The maximal height of the signal is measured by means of a digital voltmeter. The light yielder is bacterial luciferase with appropriate additives and supplemented with malate and malate dehydrogenase.The modified light-yielding solution provides for continuous formation of NADH resulting in a durable level of light emission. The cycle involved was shown not to operate with NADP⁺. The slow fading of the emission permits simplification of the measuring procedure. Rapid injection in front of the phototube can thus be omitted and replaced by ordinary mixing before the reaction cell is positioned for the measurement. Furthermore, the instrumentation required is less elaborate than in photokinetic assay, since it is not necessary to record and integrate the time course of the emission. To test the applicability of the method, analyses of pmol amounts were performed in the islets of Langerhans and in tissue samples of much smaller size than fine needle biopsies.