serial dilution Tanique

introduction

Working with billions of tiny cells can pose a problem when you need to count the total number of cells
in a sample. Fortunately, through precise serial dilution of a sample, it is possible to get down to a
number that is much easier to work with.

Serial Dilution

The answer is through dilution. If you simply pull out a smaller, exact quantity of culture liquid, you
could count those bacteria and, based on how much you pulled out of the total, you can determine how
many bacteria are in your original sample.

A serial dilution is a series of sequential dilutions used to reduce a dense culture of cells to a more
usable concentration. Each dilution will reduce the concentration of bacteria by a specific amount. So,
by calculating the total dilution over the entire series, it is possible to know how many bacteria you
started with. The best way to fully grasp serial dilutions is to try out the procedure yourself.

Manual section:

In this lab you will do the basics of serial dilution and how to use it to determine the number of cells in
an original bacterial culture in water sample

Materials for Dilutions

 sterile 1.5 ml tubes, pipet tips and micropipettors

 sterile dilution medium(buffered pepton water)

 sterile Petri plates with growth medium

 sample of water known volume

what is the dilution medium(pepton water)

Peptone Salt solution is recommended as a diluent for dilution of sample by different test methods

widely used for examination of foodstuffs. ... It contains peptone at low concentration which provides
nutrients for survival of microorganisms and hence protecting the organisms.
 Peptone Water contains peptone as a source of carbon, nitrogen, vitamins and minerals. Sodium
chloride maintains the osmotic balance of the medium.

How to Perform a Serial Dilution

I'm going to walk you through an example serial dilution using the easiest method,

To start, we need 10 milliliters (10 ml) of your original bacterial culture (labeled OBC). Before we start
diluting, we need to prepare several dilution blanks, which are tubes containing your diluting liquid in
exact quantities. Your liquid could be growth media(buffered pepton water), saline, sterile water, or any
other appropriate liquid. For this example, we need 5 dilution blanks, numbered 1-5. In each tube, we
need exactly 9 ml of liquid media. .

The tubes should be lined up like this:

How to line up tubes for serial dilution example

The first step is to gently shake or swirl the tube. This will ensure that your cells are evenly distributed in
the tube. If your cells settle to the bottom, and you remove liquid without swirling, you run the risk of
not getting enough cells, invalidating your final count. Remember to always swirl the tube before
removing liquid.

Once swirled, carefully transfer exactly 1 ml from your OBC Tube to Tube 1. Now, you should have 10 ml
of liquid in Tube 1. Exactly one-tenth of your cells are now in a new tube with a final volume of 10 ml.
You just performed a 1 in 10 dilution, or it could be written 1/10. 1 is the volume you transferred, and 10
is the final volume of the tube after thansfer.

A single dilution is calculated as follows:

 
Dilution =          volume of the sample               

volume of the sample + diluent volume

For example the dilution of 1 mL into 9 mL equals:

     1    which is the same as     1    which is written 1/10 or 10-1

    1+9                            10

This can be called, �a one to ten dilution.�

When doing very high dilutions (like 1/10,000 or 1/1,000,000), it is more accurate to do the dilution in a
series of smaller dilutions rather than in one giant dilution.  This is called a dilution series or a serial
dilution.

In a serial dilution, the final total dilution is a product of each individual dilution in the series. Thus, a
series of 5, �one to ten dilutions equals �a one to one hundred thousand� dilution:

     (1/10) x (1/10) x (1/10) x (1/10) x (1/10) = 1/100,000 = 10 -5 dilution