Silver stain for flagella Bactria Escherichia coli Pseudomonas

Silver stain for flagella • Bactria: – Escherichia coli – Pseudomonas aeruginosa Chapter 15, p 89

objective • Stain bacterial flagella which are responsible for bacterial motility

principle • Use mordant (stain solution A) to enclose flagella • Use sliver stain (stain solution B) to dye the protein of flagella

• Stain agents • Solution one (sliver stain A): – 25 m. L of saturated aluminum potassium sulfate (KAl(SO 4)2 ) – 50 m. L of 10% tannic acid – 5 m. L of 5% ferric chloride (Fe. Cl 3) • Solution two (sliver stain B): – Ammonium hydroxide (Al. OH 3) – 5% sliver nitrate

procedures • Clean slide • Drop water on the slide • Pick up bacterial colony and spread into the drop of water • Spread bacterial suspension • Air dry (DO NOT use flame) • Add solution ONE • Stain for 4 min • Wash slightly

• • Add solution TWO Stain for 2 min Place slide on an alcohol burner Do not overheat Heat the slide until steam (about 4 min) Wash Dry Observe under microscope

Pseudomonas aeruginosa

Escherichia coli

Bacillus subtilis

Motility test • Bactria: – Escherichia coli – Staphylococcus aureus • Medium – TSB + 0. 4% agar (4 g/L)(2)

• Procedure – Stab (about 1 cm) into medium – Incubate at 37℃ for 24 hours