Overview
This lecture explains the process of preparing a bacterial smear on a microscope slide, highlighting each step's importance for accurate staining and identification in microbiology.
Purpose of Smear Preparation
- A smear is a thin layer of microorganisms placed on a slide.
- Smears are made to stain microorganisms for identification and diagnostic staining.
- Staining helps determine the organism's type and whether it is potentially dangerous.
Tools and Materials
- The wire loop is the main tool used to transfer and sterilize microorganisms.
- Removing the loop leaves a needle, used for stabbing techniques.
Steps for Smear Preparation
- Sterilize the wire loop by flaming it until it is red hot.
- Let the loop cool before using it to avoid killing the organisms or causing a sizzle.
- Transfer two loopfuls of organisms onto the center of the slide.
- For liquid cultures, directly transfer two loopfuls to the slide and spread thinly.
- For solid cultures, add two loopfuls of water first, then a small amount of bacteria, and mix.
- Spread the sample into a thin layer to ensure proper staining.
Drying and Heat Fixing
- Allow the smear to air dry completely before heat fixing.
- Air drying ensures organisms adhere to the slide and prevents cell destruction during heat fixing.
- Heat fix the dried slide by quickly passing it through a Bunsen burner flame six times (about one second each pass) using a clothespin.
- Improper drying or heat fixing can damage cells and affect staining results.
Next Steps
- After heat fixing, the smear is ready for staining (e.g., Gram stain) to observe and identify organisms.
Key Terms & Definitions
- Smear — a thin layer of microorganisms placed on a microscope slide for examination.
- Wire loop — a laboratory tool used to transfer and spread microorganisms; sterilized by flaming before use.
- Aseptic technique — procedures used to prevent contamination by unwanted microorganisms.
- Heat fixing — passing a dried smear through a flame to adhere cells to the slide and kill them without destroying cell structure.
Action Items / Next Steps
- Prepare a smear using the described technique.
- Allow the smear to air dry fully before heat fixing.
- Practice proper heat fixing and staining methods as demonstrated in the lab.