Biomedical subjects

Jeffrey D Brewster

Publications and source records attributed to Jeffrey D Brewster.

3 recordsLinked to original sources

PubMed2004

Single-chain Fv antibody with specificity for Listeria monocytogenes.

Single chain antibodies (scFv) exhibiting specific binding to Listeria monocytogenes strains were isolated from a pool of random scFvs expressed on the surface of filamentous bacteriophages. Positive selection (panning) using L. monocytogenes was used to enrich for phage clones with the desired binding affinity, and negative selection using L. innocua and L. ivanovii was used to remove phages expressing cross-reactive antibody fragments. A single phage clone, P4:A8, was selected using two independent panning schemes. A rapid assay was devised to determine phage antibody binding specificity and was used to develop a selectivity profile for individual phage clones. The P4:A8 clone was screened against a panel of bacteria consisting of eight strains of L. monocytogenes, one each of the other six species of Listeria and nine other relevant bacterial species. A collection of individual clones from the penultimate panning was also screened against a subset of the panel of bacteria. The selectivity profiles indicate that multiple clones, including P4:A8, exhibit binding to one or more strains of L. monocytogenes without cross-reactivity toward any other species in the panel. This is the first report of a species-specific antibody for viable cells of L. monocytogenes (i.e., the ability to bind to L. monocytogenes without cross-reactivity toward any other species of Listeria).

Antibodies, Bacterial↗

PubMed2003

A simple micro-growth assay for enumerating bacteria.

A simple method for nonspecific determination of bacteria concentrations in a variety of liquid samples was developed. The assay was based on the time required for a sample grown in liquid media to reach a threshold turbidity. Samples were combined with media in a covered 96-well microwell plate and the turbidity was monitored in real time as the bacteria grew in a temperature-controlled plate reader. A significant problem with growth in microwells was condensation on the cover, which prevented accurate turbidity measurement. This problem was overcome by coating the cover with a small amount of surface-active agent. Salmonella and E. coli concentrations could be determined with a relative error of approximately 20% at levels from 10 to 10(6) cells/ml (eight replicates). An assay of 10 samples with standards required 10 min to set up and 20 min for data processing using a computer spreadsheet program. Growth time at 37 degrees C ranged from 4 h for samples at 10(7) cells/ml to 16 h for samples at 10 cells/ml.

Algorithms↗

PubMed2003

Isolation and concentration of Salmonellae with an immunoaffinity column.

A method for rapidly and selectively isolating Salmonellae from buffer solutions and concentrating the bacteria by a factor of approximately 500 was developed. Anti-Salmonellae antibody was covalently linked to 40 microm polyacrylamide beads to prepare a solid phase with affinity for the bacteria. The beads were packed into 1-mm diameter glass tubes to form a column 20 microl in volume. Buffer containing Salmonellae at concentrations ranging from 10(2) to 10(6)/ml was pumped through the column to trap and concentrate the bacteria. At a flow rate of 50 microl/min, more than 95% of the bacteria introduced to the column were captured, while at 800 microl/min capture dropped to 32%. Specificity was high, with no detectable capture of Escherichia coli at a concentration of 10(5)/ml. Capture of more than 90% of Salmonellae in a 5-ml sample was achieved in 40 min by re-circulating the sample through the column at a flow rate of 500 microl/.

Bacteriological Techniques↗