Assay for the identification of microorganisms
A microbial and electrochemical technology, applied in the field of analysis for identifying microorganisms, which can solve problems such as unreliability and time-consuming
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Embodiment 1
[0028] Respiratory cycle activity in the presence of effector compounds
[0029] In this example, the assay uses the following steps: (1) The method involves mixing 200 μL of bacterial culture with 1300 μL of buffer (with or without effector and 5 μM DCIP) at a fixed temperature (35° C.) The electrode was mixed in the electrochemical cell for a fixed time (10 min); (2) followed by the addition of mediator (500 μL of 0.2M potassium ferricyanide) at a fixed temperature (35°C) for a fixed time (10 min); and (3) The current is measured at a fixed voltage difference (-100mV) and for a fixed time (120sec). Ferricyanide solution was added such that the final concentration of ferricyanide was 50 mM.
[0030] In this configuration, the analysis uses approximately 0.017cm each 2 surface area of the two platinum electrodes. The measured current is proportional to the concentration of the least concentrated redox pair species in solution. In the present system, there is a large exce...
Embodiment 2
[0042] Distinguish between Gram-positive and Gram-negative bacteria
[0043] The following steps were used for each experiment: (1) Preheat 150 μL aliquots of buffer containing 1 mM glucose (glc) with or without 5 μM DCIP at a fixed temperature (35° C.) for a fixed time (4 min); (2 ) then add 50 μL of bacterial suspension to the sample at a fixed temperature (35°C) for a fixed time (10 min); (3) then add mediator (50 μL 0.4M ferricyanide Potassium) for a fixed time (10 min); and (4) current was measured at a fixed temperature (35° C.) and for a fixed time (120 sec) at a fixed voltage difference (100 mV) between two platinum electrodes. The final ferricyanide concentration in the sample prior to reaction with the microorganism was 40 mM.
[0044] In this configuration, the analysis uses approximately 0.03cm each 2 surface area of the two platinum electrodes. The measured current depends on the concentration of the least concentrated redox species in solution. In our syste...
Embodiment 3
[0051] Discrimination of 10 microbial strains using pattern recognition by principal component analysis of respiratory activity data
[0052] In this example, individual microorganisms were subjected to 22 different effector compounds in addition to control measurements. The following steps were used for each experiment: (1) Preheat 150 μL aliquots of buffer containing the effector compound and 5 μM DCIP at a fixed temperature (35° C.) for a fixed time (4 min); (2) subsequently at a fixed temperature ( Add 50 μL of bacterial suspension to the sample at 35°C) for a fixed time (10 min); (3) then add mediator (50 μL 0.4M potassium ferricyanide) at a fixed temperature (35°C) for a fixed time (10 min); and (4) current was measured at a fixed temperature (35° C.) and for a fixed time (120 sec) at a fixed voltage difference (100 mV) between two platinum electrodes. The final ferricyanide concentration in the sample prior to reaction with the microorganism was 40 mM. For each microo...
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