Sterile liquid handling is essential in microbiology, cell culture, pharmaceutical preparation, and diagnostic laboratories. The goal is to transfer liquids without introducing contamination, ensuring accuracy, reproducibility, and sample integrity. Below is a step‑by‑step guide covering sterile pipetting and dispensing techniques.
Step 1: Prepare a sterile workspace
- Wipe the bench with 70% ethanol or an approved disinfectant
- Ensure the laminar flow hood is running for at least 10–15 minutes
- Arrange sterile consumables (tips, tubes, media) within easy reach
Note: A clean environment reduces the risk of contamination before pipetting begins.
Step 2: Select the correct pipette and sterile tips
- Use micropipettes appropriate for the volume (e.g., P20, P200, P1000)
- Always use sterile, filtered tips for biological work
- Check pipette calibration and cleanliness before use
Note: Choosing the right pipette and tip ensures accuracy and prevents cross‑contamination.
Step 3: Attach the tip without contamination
- Pick up the pipette and press firmly into the tip without touching the tip with your hands
- Keep the tip away from surfaces and clothing
- Avoid reusing tips — one tip per sample
Note: Proper tip attachment prevents contamination from hands or surfaces.
Step 4: Aspirate liquid using sterile technique
- Press the plunger to the first stop before entering the liquid
- Immerse the tip 2–3 mm below the surface to avoid bubbles
- Slowly release the plunger to aspirate
- Keep the pipette vertical for accurate volume
Note: Correct aspiration ensures accuracy and prevents aerosol formation.
Step 5: Dispense liquid safely and accurately
- Touch the tip gently to the tube wall to avoid splashing
- Press the plunger to the first stop to dispense
- Press to the second stop to expel remaining liquid
- Withdraw the tip while still pressing the plunger
Note: Proper dispensing prevents contamination and ensures precise delivery.
Step 6: Dispose of tips correctly
- Eject tips into a biohazard waste container
- Never touch used tips with your hands
- Replace tips after every sample to maintain sterility
Note: Used tips must be discarded safely to prevent contamination.
Step 7: Handle reagents and media sterilely
- Open tubes and bottles only inside the laminar flow hood
- Avoid placing caps on the bench — hold them or place them facing down
- Flame sterilise glass pipettes when required (microbiology)
Note: Sterile handling prevents contamination of solutions and cultures.
Step 8: Store and label samples properly
- Label tubes clearly with sample name, date, and volume
- Store samples at appropriate temperatures (e.g., 4°C, –20°C, –80°C)
- Keep tubes closed tightly to prevent contamination
Note: Correct storage maintains sample integrity after pipetting.
Step 9: Clean up and maintain sterility
- Disinfect surfaces with 70% ethanol after finishing
- Dispose of all waste according to lab protocols
- Wash hands thoroughly after completing the procedure
Note: Proper cleanup ensures the workspace remains safe for future work.
Sterile pipetting and dispensing require careful preparation, correct equipment, and disciplined technique. By following these steps, you ensure accurate measurements, prevent contamination, and maintain the integrity of your biological work.
