Gram staining

One of the basic stains in microbiology, gram staining gave rise to the division of bacteria into Gram positive (G+) and Gram negative (G-). This division is based on the different structure of the bacterial wall.

Procedure[edit | edit source]

The sample or bacterial culture to be examined is applied to the slide and the staining solutions are gradually applied. Allow each solution to work for approximately 1 minute^[1]. The procedure is easily remembered by the abbreviation VLAS (VLAK):

Gram coloring

Positivity and negativity[edit | edit source]

G+[edit | edit source]

Gram positive

Gram-positive bacteria have a wall composed of peptidoglycan and polysaccharides through which teichoic acid passes. During staining, the crystalline violet enters the cells and forms a blue complex color with Lugol's solution. The alcohol is unable to penetrate the cell wall and dissolve the complex. Safranin staining gives the bacteria a deep purple color.

G+ cocci: Staphylococcus, Streptococcus, Enterococcus;
G+ bacilli: Corynebacterium, Clostridium, Listeria, Bacillus.

G–[edit | edit source]

Gram negative

Gram-negative bacteria have a wall consisting of a thin layer of peptidoglycan and a layer of lipopolysaccharide. In the same process, the third step involves washing out of the complex with alcohol and decolorization. Safranin stains the bacteria red.

G- cocci: Neisseria;
G- coccobacilli: Haemophilus influenzae, Bordetella pertussis, Legionella, Brucella, etc.
G- bacilli: Klebsiella, E. coli, Enterobacter, Citrobacter, Serratia, Vibrio, Pseudomonas, Proteus, Helicobacter pylori, Yersinia, Campylobacter, Salmonella, Bacillus fragilis, etc.

G labile and non-staining[edit | edit source]

Some bacteria, especially after long cultivation and multiple passaging, or if they survive the attack of antibiotics against the cell wall (L-forms), can change from G+ to G-.

Bacteria that contain a lot of fatty acids and waxes in their wall (Mycobacterium tuberculosis) can not be stained by Gram at all.