Saturday, January 22, 2011

Serial Dilutions

***UPDATE- View the updated illustration at

Many modern biology and chemistry laboratory procedures require compounds to be mixed or dissolved in a liquid such as water.  These mixtures are called solutions.  Solutions have two components: a solute and a solvent.  Solutes are the suspended or dissolved material and the liquid is the solvent.  The amount of solute present in a particular volume of water or other solvent is called its concentration. For chemicals, common concentration units include weight/volume percent, weight percent, molarity and normality.  For biologically active compounds such as antibodies or enzymes, concentrations are often described using activity units/ml.  Concentration of bacterial and cell cultures are often reported in cells / ml or CFU (colony forming units)/ml.

Many procedures require solutions at specific concentrations.  Solutions with known concentrations are prepared using one of two methods.
  1. Dissolving a known mass of a solid into water (or other solute) to a specific volume.
  2. Diluting a solution of known concentration with additional solvent.  
In addition solutions of know concentrations, many procedures require set of solutions over a range of concentrations.  Such assays include quantifying the number of bacteria in a sample of using plate counts; development of standard curves and quantitative colorimetric, radiometric and enzymatic assays. Sets of solutions over a range of concentrations are prepared via serial dilution. 

To perform a serial dilution, a small amount of a well-mixed solution is transferred into a new container and additional water or other solute is added. This dilutes the original solution.  It is important to note that the addition of more solute to a solution does not change the number of molecules or bacterial cells present. What changes is the density of the material relative to the amount of solvent. 

The initial concentration and target range needed for a given assay determines the size and number of dilution steps required. Often, serial dilutions are performed in steps of 10 or 100. They are described as ratios of the original and final concentrations.  For example a 1:10 dilution is a mixture of one part of a solution and nine parts of the solute. To make a 1:100 dilution, one part of the solution is mixed with 99 parts of the solute. For example, mixing 100 µL of a stock solution with 900 µL of water makes a 1:10 dilution. The final volume of the diluted sample would be 1000 µL or 1 mL and the concentration is be 1/10 that of the original solution.  This is also commonly referred to as a 10x dilution.

It is not uncommon for a scientist to have only a rough idea of the concentration of the solution prior to starting an assay and many assays are only effective within certain concentration ranges.  In these situations, scientists can perform the experiment on a set of serially diluted samples. Running the assay on a small set of samples over a range of concentrations ensures that one of the samples will be within the target range and give the desired result.  

The illustration below follows the relationship between volume of solvent, number of molecules of solute and concentration over a set of 4 10x dilutions. 

Demonstration video:

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