Obtaining a pure culture

The concept of the pure culture is central to classical bacteriology and is a central tenet of Koch’s postulates. If an organism can grow on agar it can be streaked to obtain single colonies each of that should have arisen as a result of a single prokaryotic cell. This works by means of the dilution effect of each round of streaking and sterilization. The 1st inoculum onto the plate might transport millions of bacteria to the agar but each time the loop is dragged across the plate these cells are removed further from their neighbors. The Coupled with sterilization of the loop fewer than 10 cells may be present in the last line of streaking before incubation.

Several prokaryotic cells will not grow on agar plates. If this is recognized to be the case then a same process of dilution is carried out in liquid broth to obtain a pure culture. One milliliter of the primary liquid culture is taken with a sterile pipette and added to 9 ml of fresh sterile medium. The 10^-1 dilution is mixed well,

Figure 1:  Diagram of the technique of obtaining single colonies through streaking. Dashed lines show the path of the loop on the surface of the agar.

Then 1 ml of which is removed and again added to 9 ml of fresh sterile medium. This procedure is repeated a further 10 or 12 times and then the incubated are dilutions. The lower dilutions will show no growth although the others will be turbid. The lowest dilution which shows growth is likely to have arisen from inoculation with less than 10 cells so repetition of this procedure will eventually lead to a pure culture. This approach can be further enhanced through making 10-fold replicates of each dilution. This should mean which the tube with growth in where less than three or four of the other replicates are growing must have arisen from a single cell. This concept can be extended into a means of generally estimating the numbers in the original culture.