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

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)

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

Mycobacterium tbc – farbenie podľa Ziehl Neelsena

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

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

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