Practical seminar 4

1. Practical seminar 4 • Acid fast staining - Mycobacterium tuberculosis • Corynebacteriuôm diphtheriae – Albert´s staining of metachromatic granules

2. Acid fast staining • Bacterial cell resistant to colorisation with some staining (Gram) • Their cell wall is relatively not permeable, containig lipids, fatty acids...(ex. Mycobacterium tbc, Corynebacteria...) • If stained (with strong basic stains (basic fuchsin in 5% fenole – they resist to decolorisation with strong acids (20% H2SO4)

3. Ziehl Neelsen staining • Heat fixed smear • Apply carbol fuchsin (basic fuchsin + phenol) • Warm untill evaporation • Rinse with water • Apply 3% HCl (or 95% alcohol • Counterstain with malachit green

4. Mycobacterium tbc – farbenie podľa Ziehl Neelsena

6. Metachromatic granules staining • Volutin storage granules in cytoplasma • Stained with basic stain • Toluidin blue, methylen blue • They are acidoresistent • Differentiation of C. difteriae from saprophytic corynebacteria • Albert´s staining

8. Metachromatic staining - principals • Stain aplied on fixed cell or tissue is of different color as is the result color • Ernst Babes inclusions – metachromatic granuls – volutine granuls – phosphate molecules - Corynebacterium diphteriae • Volutin – nucleoprotein complex – in cytoplasm of some bacteria, molds and parasits in the forme of storage granules

9. Albert´s staining • Fix smear • Apply Albert´s solution • Add Lugol solution • Bacteria are green or blue, (malachit green or methylen blue), granuls are red