Overview
This lecture explains the concept and procedure of serial dilutions in microbiology, including purpose, calculation methods, and practical steps.
Serial Dilution Basics
- Dilution reduces the concentration of a solution, making it weaker.
- Serial (log) dilutions decrease a bacterial population by consistent factors, usually by tenfold each step.
- The goal is to reach a suitable concentration for specific tests or to obtain countable colonies on agar plates.
Calculating Serial Dilutions
- A single log (10-fold) dilution means the concentration is reduced by a factor of 10.
- In a 1:10 dilution, 1 ml of sample is added to 9 ml of diluent, totaling 10 ml.
- Decimal numbers can be converted to scientific notation by moving the decimal point based on the exponent (e.g., 100 becomes 1.0 × 10²).
Performing Multiple Serial Dilutions
- Multiple serial dilutions further decrease sample concentration by repeated log reductions.
- Example: Starting with 35,000 cfu/ml, three serial dilutions reduce the concentration to 35 cfu/ml.
- First dilution: 35,000 cfu/ml → 3,500 cfu/ml.
- Second: 3,500 cfu/ml → 350 cfu/ml.
- Third: 350 cfu/ml → 35 cfu/ml (target).
Step-by-Step Serial Dilution Procedure
- Label each plate with its corresponding dilution factor.
- Place one microorganism pellet into 10 ml of pre-warmed phosphate buffer using sterile forceps.
- Vortex the suspension for even mixing.
- Pipette 1 ml from the suspension into 9 ml of buffer; repeat for each dilution step.
- Hold the pipette vertically for accuracy.
- Plate 1 ml from each dilution tube onto agar plates and use a spreader for even distribution.
- Incubate plates overnight to observe colony growth.
Importance of Serial Dilutions
- Serial dilutions are necessary to achieve plates with countable colonies, as undiluted or less diluted samples may yield too many colonies to count.
Key Terms & Definitions
- Dilution — Process of reducing the concentration of a substance in a solution.
- Serial Dilution — Repeated stepwise dilution, usually by a consistent factor (e.g., 10).
- CFU (Colony-Forming Unit) — A unit used to estimate the number of viable bacteria in a sample.
- Scientific Notation — A method for expressing large or small numbers using powers of ten.
Action Items / Next Steps
- Review and practice converting decimal numbers to scientific notation.
- Practice performing serial dilutions using 1:10 ratios and calculating final concentrations.
- Prepare for in-lab demonstration or practical assessment on serial dilution techniques.