Preview

Methyl Blue Staining on Yeast

Good Essays
Open Document
Open Document
508 Words
Grammar
Grammar
Plagiarism
Plagiarism
Writing
Writing
Score
Score
Methyl Blue Staining on Yeast
The methylene blue staining procedure is used to measure yeast viability based on the assumption that the methylene blue will enter the cells and be broken down by living yeast cells that produce the enzymes which breaks down methylene blue, leaving the cells colourless. The non- viable cells do not produce this enzyme (or enzymes) and as such the methylene blue that enters the cells are undegraded causing the cells to remain coloured (the oxidized form concentrates intracellularly).
The coloured and colourless cells are then counted using a haemocytometer and the number of viable and nonviable cells determined in a given area, the result would be then used to estimate the number of cells in the original sample. This is an easy, quick and cheap method to determine the amount of viable yeast present in a sample though this is not the best method for a number of reasons.
A major reason is that methylene blue rapidly becomes toxic to the yeast and as such preparations should be examined within 10 minutes of preparation. The older the cells become, the less likely that they are to take up the methylene blue dye from solution since as the yeast age, they deposit lipid and/or sugar in their cell membrane (in the form of free sterols[predominantly ergosterol and zymosterol with minor portions of lanosterol and fecosterol] and phospholipids[phosphotidylcholine and phosphotidylethanolamine with minor portions of phosphotidylinositol, phosphotidylserine and phosphotidyl-glycerol]) as a survival mechanism to protect their internal mechanisms from the buildup of waste in the external environment. This means that cells which are not viable would not take up the dye and due to their age and not their ability to break down the methylene blue (as a result of their viability) would remain colourless and be determined to be viable (a false positive).
The test itself is also not very accurate since yeast might not be evenly distributed in the original sample and depending on the

You May Also Find These Documents Helpful

  • Good Essays

    Task 11

    • 431 Words
    • 2 Pages

    Identification of the yeast cells by the indirect system was done effortlessly while the cells in the plaque were not that easy to recognize. After comparison, the deduction is that the direct staining offers the most distinct view of the cells.…

    • 431 Words
    • 2 Pages
    Good Essays
  • Good Essays

    The purpose of this test is to see what colors are in certain colors. We use chromatography to separate them/…

    • 595 Words
    • 3 Pages
    Good Essays
  • Good Essays

    In test 1 and test 2 each solution was placed in a separate test tube with the corresponding reagent added to each as well. In test one for proteins each substance that showed a purple color it indicated a positive result for proteins. Any solution that stayed blue is to be considered negative for proteins. The Di water in test tube one resulted in a blue color once the reagent was added. In test tube 2 Evaporated milk had the reagent added for a resulting color of milky purple. In test tube 3 the egg solution the color turned purple once the reagent was added. Finally for test one in test tube 4 containing sucrose turned light blue once the reagent was added for a negative result.…

    • 497 Words
    • 2 Pages
    Good Essays
  • Powerful Essays

    Yeast Population Lab Report

    • 2220 Words
    • 10 Pages

    The abiotic factor being tested here is what effect the temperature of the yeast’s environment has on its ability or inability to reproduce efficiently. The lab tests the yeast in three separate temperature settings: a cold temperature (4ᵒC), room temperature (22ᵒC), and a hot temperature (30ᵒC). Here, the independent variable is the temperature of the yeast habitat and the dependent variable is the amount (in mL) of CO2 gas produced by the yeast.…

    • 2220 Words
    • 10 Pages
    Powerful Essays
  • Satisfactory Essays

    The Methyl Red test shows which bacteria are creating stable acids through mixed acid fermentation of glucose. This helps to identify enteric bacteria by examining how they metabolise glucose. Every enteric bacteria first produces pyruvic acid from metabolism of glucose. A methyl red positive enteric bacteria, uses the mixed acid pathway when breaking down pyruvic acid to different acids, like lactic, acetic, and formic acids.…

    • 317 Words
    • 2 Pages
    Satisfactory Essays
  • Good Essays

    Yeast Population Growth

    • 569 Words
    • 3 Pages

    As the time increases the level of absorbance increases and the % transmission decreases in the yeast population.…

    • 569 Words
    • 3 Pages
    Good Essays
  • Better Essays

    Use a pipette to obtain a small sample of culture A, no antibiotic culture, and place it into the top notch of the hemacytometer. Do the same thing with culture B, antibiotic culture, but instead place it in the lower notch. Make sure the counting chamber does not flood, if it does clean the hemacytometer and start over. Once the samples are in the hemacytometer correctly, place a cover slip over the chamber. Place the hemacytometer on the stage of a microscope and locate the first quadrant. Use the 4X objective lens to locate the quadrant, and then switch to 10X to count the yeast cells. Count the yeast cells in all 16 squares of quadrant 1, and then repeat for quadrants 2, 3, and 4. Once the number of yeast cells are all recorded for culture A, do the same thing for culture B. Calculate the total of yeast cells in all 4 quadrants for culture A and find the average. Do the same thing for culture B. If this experiment was done with multiple groups, record the averages for those cultures as well. Once all the averages have been obtained, calculate the total average of all the group’s totals. This is known as the arithmetic mean and is calculate with the following equation: X= ΣXN. The answer obtained here will be used later on for calculating other equations. The next thing to be calculated is the variance (S²), which is used to help the variability in the…

    • 1111 Words
    • 5 Pages
    Better Essays
  • Good Essays

    Gram Staining

    • 487 Words
    • 2 Pages

    Gram staining is based on the ability of bacteria cell wall to retaining the crystal violet dye or methylene blue. The cell walls for Gram positive microorganisms have a higher peptidoglycan and lower lipid content than gram negative bacteria. Bacteria cell walls are stained by the crystal violet. Iodine is then added as a mordant to form the crystal violet and iodine complex so that the dye cannot be removed easily. This step is usually called fixing the dye. However, the next treatment with a decolorizer, which is a mixed solvent of ethanol and acetone, dissolves the lipid layer from the gram negative cells. The removal of the lipid layer increases the discharge of the primary stain from the cells into the surrounding solvent. On the other hand, the solvent dries out the thicker Grampositive cell walls, which closes the pores as the cell wall shrinks during the drying out. As a result, the diffusion of the crystal violet and iodine complex is blocked, and the bacteria remain stained. The length of the decolorization is important in differentiating the gram positive bacteria from the gram negative bacteria. A prolonged exposure to the decolorizing agent will remove all the stain from both types of bacteria. Some Grampositive bacteria may lose…

    • 487 Words
    • 2 Pages
    Good Essays
  • Good Essays

    In this experiment we used 4 large test in which we placed the dry active yeast and 30ml of heated distilled water. We also used a balance to weigh and a spatula to acquire the yeast, 250ml beakers, and a 10 ml pipette to measure. Of what was measured were…

    • 814 Words
    • 4 Pages
    Good Essays
  • Good Essays

    Our hypothesis was ‘If different amounts of molasses were tested for yeast population then the 25% molasses solution would produce the most yeast because the more the amount of molasses then the more yeast would be produced’.…

    • 641 Words
    • 3 Pages
    Good Essays
  • Satisfactory Essays

    I am testing how much carbon dioxide bubbles are produced in test tube and that will determine how efficient fermentation is working in the yeast cells. My group tested the amount of carbon dioxide bubbles produced by yeast using fructose found in apple…

    • 436 Words
    • 2 Pages
    Satisfactory Essays
  • Good Essays

    Gram Stain Experiment

    • 647 Words
    • 3 Pages

    Gram Stain: The important part of this experiment is being able to determine a bacterium based on its cell wall structure. It also helps indentify if the unknown organism is a Gram positive or Gram negative. This is the initial step that must be taken before any other lab procedure may continue on to ensure the purity is present, the arrangement of the cells, and the shape the cell has. The staining of the cell starts off with using the primary stain, Crystal Violet which is a purple color, to begin with. Next, Gram Iodine is added on, which is our mordant and this causes a complexion with the primary stain, results in formation of insoluble complexes. Then, a counter stain is applied on called Safranin (pinkish/red color). In Gram positive…

    • 647 Words
    • 3 Pages
    Good Essays
  • Satisfactory Essays

    2) If the number of bacteria in the samples is high, colonies of bacteria are likely to cover the whole plate so that one cannot count any colonies at…

    • 250 Words
    • 1 Page
    Satisfactory Essays
  • Good Essays

    Gram Staining

    • 318 Words
    • 2 Pages

    Since bacteria is really small microbiologist use stains to help them see bacteria more clearly under the microscope. Many of the stains they use color the bacteria cells directly and are called direct stains. Bacteria cells have a slightly negative charge while direct stains have a slightly positive charge that helps the stain bin to the bacteria. The strength of the binding from the stains depends on the make-up of the cell wall itself.…

    • 318 Words
    • 2 Pages
    Good Essays
  • Better Essays

    Bradford assay

    • 783 Words
    • 4 Pages

    The Bradford assay is dependent on the binding of the dye Coomassie Blue G250 to protein (mainly arginine), in which the dye is equal to the protein concentration. When the protein is absent, the solution is a red-brown colour and when the protein binds, the pKa of the dye moves causing the dye to become blue. The anionic blue form of the dye, which binds to the protein, has a maximum absorbance of 590 nm whereas the assay reagent solution of red and green forms has a maximum absorbance of 470 nm and 650 nm. Due to the assay being sensitive with a range of 20 to 200 μg protein, it can be determined by the amount of dye present in the blue ionic form. This can usually be accomplished by measuring the solution at an absorbance of 595 nm.…

    • 783 Words
    • 4 Pages
    Better Essays