Device for detecting bacterial contamination and method of use
View Patent ↗A device and method for detecting the presence or absence of a prokaryotic microorganism are provided, comprising the steps of identifying a protein, such as a microbial-specific protease that characterizes the presence of a specific prokaryotic microbe and thereby provides a marker for that microbe; detecting the protease that is a marker for the presence of a specific prokaryotic microbe by cleaving a substrate when the protease is present; and signaling the presence of that protease when cleavage has occurred. More specifically, the method comprises identifying at least one outer membrane protein or a secreted protein that is unique to a particular microbial pathogen such as for example Listeria monocytogenes and that is substrate specific.
1. A method of detecting Listeria monocytogenes in a test sample, said method comprising the steps of:
a) contacting the test sample with a peptide substrate for a protease that is unique to Listeria monocytogenes , wherein said peptide substrate comprises the sequence of SEQ ID NO:1, further comprising a fluorescent or chromagenic label and;
b) determining cleavage of the labeled peptide substrate or absence of cleavage of the labeled peptide substrate based on detection of said label, wherein the detection of said label is indicative of peptide substrate cleavage, and wherein peptide substrate cleavage is indicative of the presence of Listeria monocytogenes in the test sample, and absence of cleavage of the peptide substrate is indicative of the absence of Listeria monocytogenes in the test sample.
2. The method of claim 1 , wherein the fluorescent label comprises a fluorophore and a quencher.
3. The method of claim 2 , wherein the fluorescent label is detected using a fluorimeter or a UV lamp.
4. The method of claim 1 , wherein if the chromagenically labeled peptide substrate is cleaved, a colorimetric signal is detected by a colorimeter.
5. The method of claim 1 , wherein the labeled peptide substrate is attached to a solid surface.
6. The method of claim 5 , wherein the solid surface is glass or polypropylene.