>Pure culture indicates a single bacterial species
>After formation of culture media, put into the test tube and pour into petri dish then closed in incubator for 2to3 days , Many bacteria grown "one species".
>Isolation methods are following
1-Spread Plate Method
2-Streak Plate Method
3-Pour Plate Method
4-Serial Dilution Method
1-SPREAD PLATE METHOD
>It is the best method to isolate the pure colonies of bacteria.
>In this method a mixed culture media is diluted with distilled water or saline solution (not mixed with agar media)
>NOW take 0.1ml of culture media drop and placed on the surface of agar media plate.
>Spread it with the help of glass spreader (L-shaped) on all over the surface of agar media.
>Then allow to incubate to the agar plate and after 24 hour isolated colonies observed &counted
2-STREAK PLATE METHOD
>In this method the bacteria is isolated from mixed culture media by applying streak on the agar media with the help of inoculation loop.
>The Principle of this method is STREAKING
Procedure
>Firstly inoculation loop is heated for sterilization and then streak on the surface of mixed culture
>Then it sterak on new sterile culture media
>Plate are then incubated to allow the growth of colonies of some kind of Bacteria.
3-POUR PLATE METHOD
>In this method mixed bacteria is diluted in broth medium.
>Now, 1ml of this diluted culture media is POURED in empty sterile plate.
>Then approx 15ml of diluted agar media (45-50°c) is then transfer or pour in sample containing plate.
>Plate is then SWIRL to equally distribution of culture media and then plate agar allow to solidify.
>Then plate is INCUBATED to develop bacterial colonies in both within the agar medium (subsurface colonies) & on the medium {surface colonies}
Pour plate method
4-SERIAL DILUTION PLATE METHOD
>In this method take 5-6 test tubes filled with 9ml of distilled water or saline solution.
>Now, take 1ml solution from mixed culture tube and diluted it in first test tube.
>Then take 1ml from first test tube and diluted it in second test tube,repeat this process until the all test tubes should be diluted.
>After dilute all tubes take 1ml of each tubesand poured into nutrients agar plate , Allow them to incubate for bacterial growth.
{NOTE }:- ✓As number of test tubes increases the concentration of bacteria decreases and dilution is increases..
