Streak Plate Technique Overview

Overview

This lecture explains the streak plate technique for isolating pure bacterial colonies from mixed populations, detailing preparation, inoculation steps, and result interpretation.

Equipment and Materials

• Required items: biological safety cabinet, incubator, Bunsen burner, inoculating loop, colony counter, and prepared agar culture media plates.
• Trypton soya agar (TSA) is used as a universal growth medium.

Preparation of Culture Media

• Weigh and mix agar media with distilled water, following the media container label.
• Sterilize media as per instructions: boiling, autoclaving, or microwaving as needed.
• Pour hot media into sterile petri dishes aseptically and cool to solidify.
• Incubate newly prepared plates to check for contamination; only use fresh, uncontaminated plates.

Inoculation and Streak Plate Technique

• Clean workstation inside the biological safety cabinet before starting.
• Flame the inoculating loop until red-hot to sterilize it, and let it cool.
• Label fresh agar plates with media name, test name, and date.
• Use a cooled, sterile loop to pick up bacterial culture and smear it in one corner of the plate.
• Re-flame and cool the loop before streaking in a zigzag pattern over the first quarter of the plate.
• Repeat flaming, cooling, and streaking for each successive quarter, touching previous streaks each time.
• The four-quadrant streaking method reduces bacterial density across plate quarters.
• Cover and invert the plate, then incubate at 37°C for 24 hours.

Observation of Results

• After incubation, use a colony counter to observe colony distribution.
• Highest colony density is seen in the first quarter, lowest in the fourth.
• Isolated bacterial colonies in the third and fourth quarters can be picked for pure cultures.

Key Terms & Definitions

• Streak plate technique — a method for isolating pure bacterial colonies by spreading bacteria over an agar plate in sequential streaks.
• Agar media — a gel-like substance used for growing microorganisms.
• Tryptone soya agar (TSA) — a nutrient-rich universal growth medium for bacteria.
• Colony counter — a device used to count bacterial colonies on a plate.
• Four-quadrant streaking — a pattern to dilute bacteria for isolation by streaking each quarter of the plate separately.

Action Items / Next Steps

• Practice the four-quadrant streaking method on TSA plates.
• Observe and record colony isolation results after incubation.
• Prepare and label culture media plates for future experiments.