Method and kit for identifying antimicrobial agents and effective concentrations thereof
AssigneeCobio Diagnostics Inc.
InventorsChristopher R. Cox, Andrew J. Logan
Life of this patent
Estimating the term…
Abstract
A method and kit for for determining of an effective concentration of an antimicrobial agent for inhibiting bacterial growth are provided. The method and kit involve the use of a first and second multiwell receptacle. A fluid sample containing or suspected to contain one or more microbial organisms is dispensed into the wells of the first multiwell receptacle containing a range of dilutions of an antimicrobial agent, growth medium and a bacteriophage to form assay mixtures in the wells. The assay mixtures are then transferred to the corresponding wells of the second multiwell receptacle for detection.
Claims22
A method for determining an effective concentration of an antimicrobial agent for inhibiting bacterial growth, the method comprising:
- a) transferring a sample containing living bacterial cells to a first plurality of wells of a first multiwell receptacle, the first plurality of wells containing a range of known concentrations of at least one antimicrobial agent, each well of the first plurality of wells further containing bacterial growth medium and bacteriophage capable of infecting the bacterial cells, to thereby form assay mixtures in each well of the first plurality of wells, wherein the bacteriophage is present at a concentration that is sufficiently low to not be detectable in step d);
- b) incubating the first multiwell receptacle under conditions sufficient to promote bacterial growth and infection of the bacterial cells by the bacteriophage, and cause the production of progeny bacteriophage in the assay mixtures;
- c) transferring the assay mixtures from the first plurality of wells of the first multiwell receptacle to a corresponding second plurality of wells of a second multiwell receptacle, each of the corresponding wells of the second plurality of wells containing primary anti-bacteriophage antibodies to thereby bind the progeny bacteriophage to the primary antibodies and form a primary antibody-bacteriophage complex;
- and
- d) detecting the primary antibody-bacteriophage complex in the corresponding second plurality of wells of the second multiwell receptacle, to thereby determine an effective concentration of the antimicrobial agent, wherein an effective concentration of the antimicrobial agent is a known concentration in one or more wells in the first plurality of wells that corresponds with one or more wells in the second plurality of wells in which the primary antibody-bacteriophage complex is not detected.
A kit for determining an effective concentration of an antimicrobial agent for inhibiting bacterial growth, the kit comprising:
- a) a first multiwell receptacle comprising a first plurality of wells comprising bacteriophage capable of infecting bacterial cells in a concentration that is sufficiently low to not be detectable in step (iv);
- and
- b) a second multiwell receptacle comprising a corresponding second plurality of wells comprising primary anti-bacteriophage antibodies capable of binding the bacteriophage,
- together with instructions to determine an effective concentration of an antimicrobial agent for inhibiting bacterial growth by
- (i) transferring a sample containing living bacterial cells to a first plurality of wells of the first multiwell receptacle, the first plurality of wells containing a range of known concentrations of at least one antimicrobial agent, each well of the first plurality of wells further containing bacterial growth medium, the bacteriophage in the plurality of wells being capable of infecting the bacterial cells, to thereby form assay mixtures in each of the wells of the first plurality of wells;
- (ii) incubating the first multiwell receptacle under conditions sufficient to promote bacterial growth and infection of the bacterial cells by the bacteriophage, and cause the production of progeny bacteriophage in the assay mixtures;
- (iii) transferring the assay mixtures from the first plurality of wells of the first multiwell receptacle to a corresponding second plurality of wells of the second multiwell receptacle to thereby bind the progeny bacteriophage to the primary anti-bacteriophage antibodies and form a primary antibody-bacteriophage complex;
- and
- (iv) detecting the primary antibody-bacteriophage complex to thereby determine an effective concentration of the antimicrobial agent, wherein an effective concentration of the antimicrobial agent is a known concentration in one or more wells in the first plurality of wells that corresponds with one or more wells in the second plurality of wells in which substantially no labeled primary antibody-bacteriophage complex is detected.
Dependencies are inferred from the claim text.