Method for determining ATP

A solution, a technology of benzalkonium bromide solution, applied in the field of determination of ATP content, achieves good application prospects and practical value, and the effect of improving sensitivity and accuracy

Inactive Publication Date: 2012-12-12
TONGJI UNIV
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  • Summary
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  • Application Information

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Problems solved by technology

For the ATP detection of solid samples like soil, there is currently no fast and efficient method, so it is very valuable and necessary to find a more widely applicable ATP detection method with high sensitivity and reliability

Method used

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  • Method for determining ATP
  • Method for determining ATP

Examples

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Embodiment 1

[0023] Determination of the amount of ATP in E. coli standard solution

[0024] Escherichia coli standard solution: Put single bacteria (E.coli.Top10) into 1mL LB liquid medium for overnight culture, take 1mL of cultured E. coli culture medium and transfer it to 40mL LB liquid medium, the speed is 120r min -1 , cultivated for 2h. Take 0.5mL, put the culture solution into 4.5mL of sterilized distilled water, make the E. coli culture solution with a dilution gradient of 10 times, and prepare the dilution gradient of 10 times according to this method 2 and 10 3 times of Escherichia coli culture medium. The ATP in the E. coli standard solution of each dilution factor was measured respectively.

[0025] The determination procedure of the E. coli bacterial liquid is as follows: take 1 mL of the culture liquid and quickly add 4 mL of 0.05% BAB solution, and shake for 90 seconds. Draw 100 μL of the mixed solution into a filter cuvette that has been added with 100 μL luciferase, an...

Embodiment 2

[0032] Determination of the amount of ATP in soil samples

[0033] Soil samples: 3 different types of soil were collected: the farmland in the 98 embankment of Chongming Island, Shanghai, on which cauliflower, pasture and corn were planted respectively; the soil of 2 experimental improvement plots in Chongming Dongtan Wetland Park and the control soil; Jiuduansha, Shanghai Wetland saline algae zone (light beach), sea three-edge clamgrass zone (Scirpusmariqueter) and soil of Spartina alterniflora community zone (Spartina alterniflora). Five parallel samples were taken for each of the three types of soil according to the diagonal method (surface soil 5-20cm). Fresh soil samples were stored at 4°C for the determination of soil ATP value and microbial indicators.

[0034] The detection steps for soil samples are: put 5g of fresh soil samples into 50mL of sterilized water, add 10-15 glass beads, put them in a shaking table and shake for 60min at a speed of 160r min -1 . Draw 2mL...

Embodiment 3

[0041] Determination of ATP content in sludge

[0042] Sludge source: Sludge from the aeration tank of Quyang Sewage Treatment Plant was collected on June 1, 2010 and June 24, 2010. Test steps for sludge: Dilute the sludge 20 times with sterilized distilled water to prepare a sludge solution; draw 2mL of soil sample sludge solution, quickly add 8mL of 0.05% BAB solution, and shake for 45s. Filter with quantitative filter paper, draw 100 μL of the filtrate into a filter cuvette that has been added with 100 μL luciferase, absorb and discharge the solution in the cup 2 to 3 times to mix the solution evenly, and then measure ATP.

[0043] We use the colony counting method and the method of the present invention and its Profile-1 bioluminescence instrument standard method to measure simultaneously, and compare them, and the results as described in Table 3 can be obtained.

[0044] Table 3: Concentration of microorganisms in activated sludge of Quyang sewage treatment plant

[004...

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Abstract

The invention relates to a method for determining ATP. The method is used for determining ATP content in bacteria-containing water body, soil, sludge, etc. The method comprises the following steps of a) preparing a to-be-measured solution of a sample; b) adding a BAB solution in the to-be-measured solution; c) sucking up a filtered filtrate and putting into a filtering cuvette with luciferase added therein already; and d) performing an ATP determination by using a Profile-1 biological luminescent instrument. Compared with a conventional method, the method provided by the invention is superior to a standard analysis method installed in the Profile-1 biological luminescent instrument, is particularly suitable for determining ATP in the soil, and makes up disadvantages of the standard analysis method installed in the Profile-1 biological luminescent instrument.

Description

technical field [0001] The present invention relates to a method for measuring ATP content, in particular to a method for measuring ATP content in bacteria-containing water, soil, and sludge using a Profile-1 bioluminescence instrument, and indirectly expressing microorganisms in soil by measuring ATP content Quantitative method. Background technique [0002] ATP (Adenosine Triphosphate) is a specific high-energy phosphate compound in organisms. The ATP content in normal organism cells is relatively stable. Therefore, the ratio of ATP content to active microbial mass can be considered as a constant. Therefore, ATP can be used as an indicator to characterize the active microbial biomass. The ATP bioluminescence method uses ATP to react with luciferin-luciferase to generate photons, and then detects the luminescence value with a fluorescence detector to obtain the amount of ATP. Its reaction process is as follows: [0003] [0004] [0005] At present, there are mai...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/76G01N1/28
Inventor 席雪飞胡煜张艳楠彭晓佳王磊武满满唐玉姝
Owner TONGJI UNIV
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