I have heard that people use haemocytometer to count bacteria, will this work or are the cells too small to be seen?
Hi Caroline,
You can also use OD (optical density) measurement. It s fast and easy to determine the cell concentration.
Bests,
Ilknur
Thanks very much, Ilknur.
I have tried that but it didn't quite work for my experiment. I need the cells at a low concentration of about 0.5 McFarland standard but when I read it with the spectrophotometer at 600nm, it is usually close to zero.
Hemocytometer will work. Label the strains with a live stain from Invitrogen if you need better contrast to visualize and count them. This takes only about 10 minutes if done properly. Be certain, however, your culture is actively growing before using the hemocytometer if you don't use a live stain as dead bacteria will also be present.
Caroline you can still use OD. Just work out an OD that gives you a reliable number of colony forming units per millilitre (cfu/ml). Then you can prepare subsequent microbial suspensions at that OD (for which you know the cfu/ml) and dilute them to your desired concentration. This way you can use a microbial turbidity (OD) that gives you a reliable cfu/ml and dilute to get down to the 1 x 10^8 that corresponds to 0.5 McFarland.
Meant to add that you don't have to use pour plates to determine cfu/ml. You can do something like a Miles & Misra method where multiple 10 ul drops of serial dilutions are placed on the surface of agar plates. After incubation you count the number of colonies in each of the drops of the countable serial dilution, calculate the average number of colonies and work out what your original suspension contained.
Dear Christine, thanks very much for your thoughts, and for introducing to me the Miles & Misra method! I think it's a great alternative to pour plating...
Use of heamocytometer for direct microscopic count is faster method but, it is not able to differentiate between viable and dead cell. So, it is not accurate method that can be considered as alternative to pour plate method. May be you should try OD measurement.
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