Pour Plate Technique - Principle, Formula, Procedure, Enumeration, Advantages, Disadvantage

Introduction to Pour Plate Technique

The pour plate technique of enumeration is one of the widely used procedures for the enumeration of viable microorganisms.

Principle of Pour Plate Technique

The principle of pour plate technique is based on the principle that when a single viable microbial cell that has been separated by dispersion from one another in a confined space of an agar medium will give rise to a macroscopically visible separate colony.

This determines the viable cells in the initial population of the microorganism by counting the number of colonies that grow on the inoculated medium after inoculation and multiplying that number by the dilution factor which is called viable count.

The dilution is usually done in a series of test tubes containing sterile distilled water or physiological saline which does not contain any nutrients. A measured amount of suspension is taken from diluted samples and poured onto an empty sterile Petri plate and nutrient media at 45-50°C is poured on top of the inoculum.

Formula of Pour Plate Technique

For pour plate technique, the number of organisms can be determined by using the formula:

No. of organism per ml = no. of colonies / Dilution x inoculum size

Where.

Dilution = volume of sample / total volume of sample and diluent

Procedure of Pour Plate Technique

Methods used for/procedure of pour plate technique include:

Sterile dilution method

The sterile dilution method is done by following steps:

1. Take sterile dilution blanks with each containing 9ml and mark it as 10^-1, 10^-2,......10^-n

2. Take 1ml or 1 gram from the sample with the help of a sterile pipette and mix it with the first dilution blank measured 10^-1, thus preparing an initial dilution

3. Shake gently to mix the content to obtain uniform distribution of cells

4. From the initial dilution i.e. 10^-1, take 1 ml of suspension by using a sterile pipette and mix it with 10^-2

5. Repeat the procedure until the desired dilution (10^-n) is obtained

6. From selected dilutions, pipette out 1 ml and empty sterile Petri plates

7. Pour approximately 15ml of molten (45-50°C) Na onto the inoculated petri dish

8. Immediately homogenize the mixture by gently moving the plate in a "Z" or "∞" pattern before the agar is able to solidify

Enumeration of Pour Plate Technique

The enumeration for the Pour Plate Technique is:

No. of organism per ml = no. of colonies / Dilution x 1

Advantages of Pour Plate Technique

The advantage of the Pour Plate Technique is:

• Both isolation and enumeration can be done by this method

Disadvantage of Pour Plate Technique

The disadvantage of the Pour Plate Technique includes:

• Some viable microorganisms may be trapped beneath the surface of agar medium and can not grow

• The organisms do not need to withstand the high temperature of the liquid agar thus psychrophiles are able to grow