Why Sterile Technique Matters

Sterile technique is the foundation of successful mushroom cultivation. Without proper sterile practices, contamination will overtake your cultures before mushrooms can develop. This guide teaches you the essential skills needed to work cleanly and efficiently.

Understanding Contamination Sources

Common Contaminants

• Mold spores: Present in air and on surfaces
• Bacteria: From hands, breath, and environment
• Yeasts: Often from skin and organic matter
• Viruses: Less common but possible

Contamination Vectors

1. Airborne particles
2. Direct contact
3. Non-sterile tools
4. Compromised containers
5. Poor technique

Setting Up Your Workspace

Creating a Still Air Box (SAB)

A still air box is your first line of defense against contamination:

1. Materials needed:

   • Large clear storage tote (minimum 60 quart)
   • Drill with 4-inch hole saw
   • Sandpaper
   • Duct tape for smooth edges

2. Construction:

   • Cut two armholes in the front
   • Sand edges smooth
   • Cover sharp edges with tape
   • Clean thoroughly before first use

Workspace Preparation

Before any sterile work:

1. Clean the room thoroughly
2. Turn off fans and HVAC
3. Close windows and doors
4. Allow air to settle (30 minutes)
5. Wipe all surfaces with 70% alcohol

The Sterile Technique Process

Pre-Work Preparation

1. Personal hygiene:

   • Shower before work
   • Wear clean clothes
   • Tie back long hair
   • Remove jewelry
   • Wash hands thoroughly

2. Tool sterilization:

   • Pressure cook implements at 15 PSI for 30 minutes
   • Flame sterilize metal tools
   • Prepare alcohol spray bottles

Working in the SAB

1. Setup:

   • Spray SAB interior with alcohol
   • Wipe all surfaces
   • Place materials inside
   • Put on gloves
   • Spray gloves with alcohol

2. Movement principles:

   • Move slowly and deliberately
   • Minimize hand movements
   • Keep hands low in the box
   • Never reach over open containers
   • Work from cleanest to least clean

Flame Sterilization

For metal tools like scalpels and needles:

1. Heat to red-hot in flame
2. Cool in sterile area (not by blowing!)
3. Test temperature on agar edge
4. Re-sterilize between each use

Common Mistakes to Avoid

Beginner Errors

• Working too fast
• Forgetting to sterilize between steps
• Breathing directly over work
• Touching non-sterile surfaces
• Opening too many containers at once

Environmental Mistakes

• Working in drafty areas
• Not allowing air to settle
• Using contaminated materials
• Poor storage of sterile items

Testing Your Technique

Agar Test Plates

Create test plates to evaluate your technique:

1. Pour sterile agar plates
2. Expose in different conditions
3. Incubate at room temperature
4. Check for growth after 3-5 days

Improvement Metrics

• Zero contamination = excellent
• 1-2 colonies = good, needs refinement
• 3+ colonies = review technique

Advanced Tips

Flow Hood Alternative

For serious cultivators, consider building a laminar flow hood:

• Provides consistent clean airflow
• More reliable than SAB
• Allows for larger-scale work

Positive Pressure Room

Create slight positive pressure:

• HEPA filter on intake
• Sealed room
• Maintains cleaner environment

Troubleshooting Contamination

If Contamination Occurs

1. Identify the type (color, texture, smell)
2. Review your process
3. Check all materials
4. Improve weak points
5. Start fresh with better technique

Prevention Checklist

• Workspace cleaned
• Tools sterilized
• Hands washed
• Gloves sprayed
• Moving slowly
• Flame sterilizing between uses

Next Steps

Once you’ve mastered basic sterile technique:

1. Practice with agar work
2. Try liquid culture preparation
3. Attempt grain-to-grain transfers
4. Work with more sensitive species

Remember: Good sterile technique is about consistency and attention to detail. Every contamination is a learning opportunity to improve your process.