Experiment 1: Direct Microscopic Count of Microorganisms in Milk

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Last updated 1:29 AM on 5/26/26
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14 Terms

1
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What is the microbial population of milk typicallt composed of?

  • Gram Positive

  • Non-motile

  • Microaerophilic

  • Anaerobic

  • Rods or Cocci

2
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What are some examples of genera represented by the microbial population of milk?

  • Lactobacillus

  • Micrococcus

  • Streptococcus

3
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How can the microbial content of a milk sample be determined?

  • microscopic examination of a stained film of a milk sample

4
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What are some advantages of the stained film method?

  1. Fast results

  2. Can count individuals and clumps of bacteria

  3. Minimal equipment

  4. Can determine bacterial morphology

  5. Slides kept for permanent record

5
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What are some disadvantages of the stained film method?

  1. Inaccurate counts on low-count milk

  2. Dead cells in pasteurized milk are counted = overcounting cells

  3. Continuous microscope use = tiring

  4. Can confuse extraneous debris as bacterial cells

6
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What are the types of milk we used?

  1. Raw unpasteurized milk

  2. Fresh pasteurized milk

  3. Expired Pasteurized milk kept in fridge

7
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What staining solutions did we use?

  1. Methylene blue 0.14%

  2. SlideBrite

8
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What is MF?

  • Microscope factor

  • relationship of the field to a milileter

9
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How to calculate MF?

  • use stage micrometer to measure the diameter of the oil immersion field.

  • Formula = πr² = area

10
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What to look through to measure the diameter?

  • Eyepiece

  • DO NOT use the camera image

  • Each space = 0.01mm

11
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How to calculate MF? (steps)

  1. Count # of spaces (ex. # = 16)

  2. Diameter = 16× 0.01mm = 0.16mm

  3. Apply πr²

    1. divide diameter/2 (0.16/2 = 0.08mm)

    2. π(0.08)² = 0.02mm

  4. Convert mm to cm

    1. 0.02mm / 100 = 0.0002cm²

  5. Number of fields = 1cm² / 0.0002cm² = 5000

  6. MF = # fields in 1mm

    1. = # fields in 1cm area / vol of milk over 1cm

    2. = 5000 fields/cm² / 0.005ml/cm²

    3. = 1,000,000 fields/mL

12
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Why is methylene blue used in the direct microscopic count method?

Provides distinct contrast between the organism and the backrground

13
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Direct count vs. Standard plate count

Direct method observed stained slides which provides faster results but counts both live and dead cells.

14
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Why do we need to steam fix the smear before staining in the direct count method?

Ensure that all cultures arent washed away during away.