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Name: Hoang
Status: Student
Grade:  Other
Location: N/A
Country: United States
Date: January 2006


Question:
Dear Madam/Sir,

As to an answer about the E coli in the NEWTON archives stating that after incubating for 24 hours, the culture should be turbid and contain 10^8 and 10^9 cells/ml. My question is how can we determine that number?



Replies:
You have to obtain a Petroff-Hauser counting chamber. This is a glass slide with very fine etchings and a coverslip. Whan viewed under a microscope (40X objective), each "square" represents 0.00000005 mL. After placing a drop of the bacterial culture on the slide, you count the number of bacteria in several squares (at least 100 cells) and then calculate the average number of cells per square and then multiply the average by 20,000,000 to find the number of cells/mL.

A countin Petroff-Hauser counting chamber can be purchased from VWR Scientific.

Good luck,

Ron Baker, Ph.D.


One could do a serial dilution. Since we don't know exactly how many organisms are there, we have to make many dilutions of the broth until we reach a dilution that can be counted. We dilute the broth 1/10, 1/100, 1/1000 etc. usually 6-7 times. We plate 1 ml of each dilution and incubate. We count a plate that has between 30-300 organisms. Then we multiply back times the dilution factor. So for instance, if you count 125 organisms on the 1/10,000 plate, you multiply 125 x 10,000. That is how many organisms were in the original broth.

vanhoeck



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