Method for measuring cells, and reagent for cell measurement
View Patent ↗A method and a means are provided, by which multiple types of cells can be simultaneously measured with high sensitivity by an ATP luminescence method. A method for measuring cells in a sample is provided, which comprises the steps of adding methanol to a sample suspected of containing viable cells to increase ATP within viable cells, extracting intracellular ATP, and causing extracted ATP to emit luminescence.
1. A method for measuring cells in a sample, comprising:
a first step comprising adding a solution comprising 0.1% to 20% methanol to a sample suspected of containing viable cells to increase ATP within the viable cells and adding an enzyme to remove ATP outside of the viable cells:
a second step of extracting intracellular ATP following the first step of increasing intracellular ATP and removing extracellular ATP; and
a third step of causing the extracted intracellular ATP to emit luminescence and measuring a luminescence level thereof.
2. The method according to claim 1 , wherein in the first step, a substance selected from a sugar and alanine is further added to the sample.
3. The method according to claim 2 , wherein the sugar is at least one sugar selected from the group consisting of glucose, fructose, and sucrose.
4. The method according to claim 2 , wherein the final concentration of the sugar is 0.8 mM or less.
5. The method according to claim 2 , wherein the final concentration of alanine is 0.5 mM or less.
6. The method according to claim 2 , wherein the final concentration of methanol is 1%, the final concentration of the sugar is 0.1 mM, and the final concentration of alanine is 0.1 mM.
7. The method according to claim 1 , wherein in the first step, the methanol and enzyme are added simultaneously and the enzyme is ATPase.
8. The method according to claim 1 , wherein after the first step, cells are captured by a filter via filtration with the filter, a solution containing methanol is removed, and then the second and the third steps are performed.
9. The method according to claim 1 , wherein suspected of containing viable cells has been prepared by filtering a solution suspected of containing viable cells and the first step and the second step are performed on the filter used.
10. The method according to claim 1 , wherein a microorganism of at least one genus selected from the group consisting of the genus Bacillus , the genus Pseudomonas , the genus Methylobacterium , the genus Escherichia , the genus Staphylococcus , the genus Clostridium , the genus Candida , and the genus Aspergillus is used as a positive control.
11. The method according to claim 1 , wherein at least one microbial species selected from the group consisting of Bacillus subtilis, Pseudomonas aeruginosa, Pseudomonas fluorescence, Methylobacterium extorquens, Escherichia coli, Staphylococcus aureus, Clostridium sporogenes, Candida albicans , and Aspergillus niger is used as a positive control.
12. The method according to claim 1 , wherein in the first step, the solution comprising methanol is removed prior to the second step.