AUTOMATED YEAST BUDDING MEASUREMENT
The invention generally relates to analyzing yeast viability and reproduction rate of yeasts. More particularly, the invention relates to efficient and effective methods and compositions for accessing and measuring budding percentages, viability and concentration of yeast cells.
1 . A method for automated analysis of budding status of yeast cells, comprising:
staining a sample to be analyzed for yeast cell budding with a dye in a buffer solution;
acquiring a fluorescent image of the dye-stained sample;
analyzing the fluorescent image of the dye-stained sample to determine the aspect ratio of the images of yeast cells in the dye-stained sample by a computer-based automated process, thereby determining the status of budding yeast cells in the sample.
2 . The method of claim 1 , wherein the computer-based automated process comprises automated measurement of the shape of round budding yeasts.
3 . The method of claim 2 , wherein a round yeast cell is considered budding if its aspect ratio is 1.1 or greater.
4 . The method of claim 1 , wherein the dye is selected from the group consisting of Acridine Orange, SYTO 9, DAPI, Hoechst, Calcofluor White, Propidium Iodide, Ethidium Bromide, Oxonol, Mg-ANS, Acriflavine, ConA-FITC.
5 . The method of claim 1 , wherein the buffer condition has a pH of about 5 to about 12.
6 . (canceled)
7 . The method of claim 1 , wherein the sample to be tested is a sample from a process of alcohol production using yeast.
8 . The method of claim 1 , wherein the sample to be tested is a sample from a biofuel fermentation process, a wine production process or a beer brewing production process.
9 . (canceled)
10 . The method of claim 8 , wherein the sample to be tested comprises one or more debris of corn mash, sugar cane, cellulose and corn stover.
11 - 15 . (canceled)
16 . The method of claim 1 , wherein the yeast is the species of Saccharomyces cerevisiae.
17 . The method of claim 4 , wherein the dye is Acridine Orange having a concentration of about 1 μg/mL to about 50 μg/mL.
18 . A method for simultaneously determining yeast budding and viability, comprising:
staining a sample to be tested with a first dye and with a second dye in a buffer solution;
acquiring a first fluorescent image of the sample stained with the first and second dyes, the first fluorescent image corresponding to the fluorescence from the first dye;
acquiring a second fluorescent image of the sample stained with the first and second dyes, the second fluorescent image corresponding to the fluorescence from the second dye;
analyzing the first fluorescent image to determine the aspect ratio of yeast cells by a computer-based automated process, thereby determining the status of budding yeast cells in the sample; and
analyzing the second fluorescent image to determine yeast viability.
19 . The method of claim 18 , wherein the computer-based automated process comprises image analysis to measure the shape of budding yeasts.
20 . The method of claim 18 , wherein an image of yeast cell is considered budding if its aspect ratio is 1.1 or greater.
21 . The method of claim 18 , wherein the first dye is selected from the group consisting of Acridine Orange, SYTO 9, DAPI, Hoechst, Calcofluor White and the second dye is selected from the group consisting of Propidium Iodide, Ethidium Bromide, Oxonol, Mg-ANS.
22 . The method of claim 21 , wherein the first dye is Acridine Orange and the second dye is Propidium Iodide.
23 . The method of claim 18 , wherein the buffer condition has a pH of about 5 to about 12
24 . (canceled)
25 . The method of claim 18 , wherein the sample to be tested is a sample from a biofuel fermentation process, a wine production process or a beer brewing production process.
26 . The method of claim 25 , wherein the biofuel fermentation process comprises producing ethanol, butanol or methanol.
27 - 28 . (canceled)
29 . The method of claim 25 , wherein the sample to be tested comprises debris of corn mash, sugar cane, cellulose or corn stover.
30 - 32 . (canceled)
33 . The method of claim 18 , wherein the yeast is the species of Saccharomyces cerevisiae.
34 . The method of claim 22 , wherein Acridine Orange is at a concentration of about 1 μg/mL to about 50 μg/mL and Propidium Iodide is at a concentration of about 1 μg/mL to about 50 μg/mL.
35 . The method of claim 18 , further comprising analyzing the first and second fluorescent images to determine concentration of budding yeast cell.
36 . A method for simultaneously measuring concentration, viability, budding percentage of yeast cells in a sample, comprising:
staining a sample to be tested with a first dye and with a second dye under a buffer condition having a pH of about 5 to about 12;
acquiring a first fluorescent image of the sample stained with the first and second dyes, the first fluorescent image corresponding to the fluorescence from the first dye;
acquiring a second fluorescent image of the sample stained with the first and second dyes, the second fluorescent image corresponding to the fluorescence from the second dye; and
analyzing the first and second fluorescent images to determine the concentration, viability, and budding percentage of yeast cells.
37 . The method of claim 36 , wherein the first dye is selected from the group consisting of Acridine Orange, SYTO 9, DAPI, Hoechst, Calcofluor White and the second dye is selected from the group consisting of Propidium Iodide, Ethidium Bromide, Oxonol, Mg-ANS.
38 . The method of claim 37 , wherein the first dye is Acridine Orange and the second dye is Propidium Iodide.
39 . The method of claim 36 , wherein the buffer condition has a pH of about 8 to about 12.
40 . (canceled)
41 . The method of claim 36 , wherein the sample is from a biofuel fermentation process, a wine production process or a beer brewing production process.
42 . (canceled)
43 . The method of claim 41 , wherein the sample comprises one or more debris of corn mash, sugar cane, cellulose and corn stover.
44 . (canceled)
45 . (canceled)
46 . The method of claim 36 , wherein the yeast is the species of Saccharomyces cerevisiae.