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A O Ruggles

Publications and source records attributed to A O Ruggles.

5 recordsLinked to original sources
PubMed1996

Chicken anti-protein A prevents Staphylococcus aureus protein A from binding to human and rabbit IgG in immunoassays and eliminates most false positive results.

This report demonstrates that chicken anti-protein A can prevent both soluble and surface-bound Staphylococcal protein A from binding to either human or rabbit IgG. In an ELISA assay, chicken anti-protein A prevented > 98% of the soluble protein A from binding to the human IgG-Fc coat. In a blotting assay, chicken anti-protein A prevented the membrane-bound protein A from interacting with the human IgG probe. When intact S. aureus (Cowan I strain) was bound to the surface of a microassay plate, chicken anti-protein A blocked > 98% of the cell wall protein A and permitted the probing of the surface components with human IgG. In another immunoassay, rabbit anti-enterotoxin A IgG was used to measure enterotoxin A concentrations in S. aureus culture medium supernatants after soluble protein A was blocked by chicken anti-protein A. Thus, the binding of chicken anti-protein A to protein A almost completely eliminates false positive results and permits the measurement of specific antibodies or antigens in a variety immunoassays where protein A is present.

Animals↗
PubMed1993

Antibodies bound to nitrocellulose in acidic buffers retain biological activity.

This report compares the binding of proteins to nitrocellulose membranes in acidic buffers (pH 2 and 3) with binding in neutral buffer (pH 7). Initially, similar amounts of antibodies and other proteins bound to the nitrocellulose membrane in both acidic and neutral buffers. However, the susceptibility of individual proteins to displacement (stripping) from the membrane by the milk blocking agent depended on the pH of the buffer used to bind the proteins to the membrane. Most proteins that bound to nitrocellulose in acidic buffers were relatively resistant to milk-stripping compared to proteins bound in pH 7 buffer. Acid-binding of proteins to nitrocellulose also decreased the amount of protein that was stripped from the nitrocellulose membrane when Tween 20 was included in the washing buffer. After correcting for the amount of antibody remaining on the membrane after the milk block, it was found that acid-bound antibodies were unchanged in biological activity when compared with the same antibodies bound at neutral pH. These results suggest that acid-binding of proteins could increase the sensitivity of nitrocellulose membrane assays that use milk and/or Tween 20.

Animals↗
PubMed1992

Characterization of immune complex components by dot blot analysis.

A method is described for the characterization of immune complex components by dot blot analysis. After isolation by chromatographic techniques and precipitation with polyethylene glycol, immune complexes were dissociated in 0.1 M phosphate (pH 2) and bound to a nitrocellulose membrane in a dot blot unit. Biotinylated probes were then used to identify the following immune complex components: specific antigens, biologically active antibodies, antibody isotypes, antibody subclasses, antibody idiotypes, and rheumatoid factors. This nonradioactive procedure takes less than 2 h to perform and has been used to analyze immune complexes isolated from sera (rabbit and human) and synovial fluid (human).

Acids↗
PubMed1991

Binding of antibodies and other proteins to nitrocellulose in acidic, basic, and chaotropic buffers.

This report compares the binding of proteins to nitrocellulose membranes in acidic buffers (pH 2 and 3) with binding in neutral buffer (pH 7), basic buffers (pH 12 and 13), 8 M urea (pH 2, 3, and 7), and 6 M guanidine hydrochloride (pH unadjusted). Initially, similar amounts of antibodies and other proteins bound to the nitrocellulose membrane in all of these buffers and solvents. However, the susceptibility of individual proteins to displacement (stripping) from the membrane by the milk blocking agent depended on both the pH and the type of buffer or solvent used to bind the proteins to the membrane. Most proteins that were bound to nitrocellulose in acidic buffers were relatively resistant to milk stripping compared to proteins bound in pH 7 buffer. After correction for the amount of antibody remaining on the membrane after the milk block, it was found that acid-bound antibodies were unchanged in biological activity when compared with the same antibodies bound at neutral pH. These results suggest that acid binding of proteins could increase the sensitivity of nitrocellulose membrane assays using a milk block.

Animals↗