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Measuring Microbial Growth

Measuring Cell Growth. Number of Cells per milliliter of liquidcells/mlBacterial cultures contain millions of cellsDirect Vs. indirect countsDirect Counts mean actually counting cellsMore accurate, but takes a lot longerIndirect Counts mean estimating the number of cells based on turbidity in

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Measuring Microbial Growth

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    1. Measuring Microbial Growth Lab 5

    2. Measuring Cell Growth Number of Cells per milliliter of liquid cells/ml Bacterial cultures contain millions of cells Direct Vs. indirect counts Direct Counts mean actually counting cells More accurate, but takes a lot longer Indirect Counts mean estimating the number of cells based on turbidity in liquid or dry weight. Less accurate, but quick Total Count vs. Viable count Total count?all cells in a culture Viable count?only live cells

    3. Turbidity Method Spectrophotometer Measure % transmittance Amount of light going through culture Transmittance is inversely proportional to turbidity (cell concentration) As turbidity increases cell number increases and transmittance decreases Indirect and Total Count

    4. Turbidity and Transmittance

    5. LEARN YOUR SCIENTIFIC NOTATION BY THURSDAY http://a-s.clayton.edu/furlong/BIOL2250LAB/mediaguide/Scienctific%20Notation%20Help.doc Or there is a link on the course web.

    6. Plate Counts Direct Viable counts The idea is to evenly spread the culture across a plate and count the colonies Each colony represents a cell in the original culture. Problem: If we have a culture that has 1x106 cells/ml and we put 1 ml on the plate will we have isolated colonies?

    7. Plate Count We must dilute first If we put 1ml of culture into 9 ml of sterile media we have performed a 1/10 serial dilution (SD) Formula: SD=AT/TV AT=amount of culture transferred TV=Total volume after transfer (1ml + 9 ml) What if we did 5 serial dilutions on that culture that had 1 x 106 cells/ml

    9. Now if we have 1 x 106 cells/ml in the original culture and we diluted it by 1/100,000 or 1 x 10-5 (as in Tube 5) then how many cells per ml do we have in tube 5? (1 x 106 cells/ml) X (1 x 10-5 ) = 1 x 101= 10 cells/ml Now we must plate it out!

    10. Practice I have a pond water with 1 x 108 cells/ml I do the following dilution. Transfer 1ml from pond water to A which has 9 ml of sterile media. Transfer 0.1 ml from A to B which has 9.9 ml of sterile media. Transfer 0.1 ml from B to plate. How many colonies should I expect to see on the plate?

    11. The Reverse Rarely do we actually know the concentration of the starting culture. Our point is to find this out! Let’s look at this dilution scheme.

    13. PRACTICE You have practice problems on the web! Practice problems on the objectives! Practice problems on the data sheet!

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