Gram stain
Gram-positive bacteria are those that are stained dark blue or violet by the staining process. Organisms that are Gram-positive have cell walls containing multiple layers of peptidoglycan bound together by amino acid bridges.
Technique
Not all bacteria reliably take either stain. Mycobacteria, for example, tend to need the Ziehl-Neelsen stain.
- Prepare a heat-fixed smear of the material containing the bacteria of interest
- Stain with crystal violet
- Treat with iodine solution, usually Lugol's solution. Its mordant solution fixes the crystal violet in Gram-positive cell walls.
- Decolorize with an alcohol or acetone-alcohol
- Counterstain with safranin, a red dye
Clinical correlations
Broad-spectrum antibiotics affect both Gram-positive and Gram-negative bacteria. No antibiotic attacks all Gram-positive, all Gram-negative, or any other total population of bacteria. Gram staining is a guide to antibiotic selection, but bacterial culture and antibiotic sensitivity testing as well as clinical experience — especially local to a given area — remain the gold standard of selectng antibiotics for treatment.
Gram-positive
Especially when antibiotics were new, a given antibiotic, such as penicillin , of the beta-lactam class, tended to attack only Gram-positive bacteria.
Gram-negative
Other classes, such as streptomycin, a member of the aminoglycoside class, tended to attack Gram-negative bacteria.
Broad-spectrum
Broad-spectrum antibiotics attack both Gram positive and negative bacteria.