Put the following substances in the order they are used in a Gram stain: counterstain, decolorizing agent, mordant, primary stain.

Verified step by step guidance

1. Start by identifying the purpose of each substance in the Gram staining process.
2. The primary stain is used first to color all cells.
3. The mordant is applied next to fix the primary stain to the cell walls.
4. The decolorizing agent is then used to differentiate between Gram-positive and Gram-negative cells.
5. Finally, the counterstain is applied to provide contrast to the decolorized cells.

Key Concepts

Gram Staining Procedure

Gram staining is a differential staining technique used to classify bacteria into two groups: Gram-positive and Gram-negative. The process involves a series of steps that include applying a primary stain, a mordant, a decolorizing agent, and a counterstain. Understanding the sequence of these steps is crucial for accurately identifying bacterial cell wall characteristics.

Primary Stain

The primary stain, typically crystal violet, is the first dye applied in the Gram staining process. It penetrates the bacterial cell wall and stains all cells. This step is essential as it provides the initial color that differentiates between Gram-positive and Gram-negative bacteria based on their cell wall structure.

Decolorizing Agent

The decolorizing agent, usually ethanol or acetone, is applied after the primary stain and mordant. It removes the primary stain from Gram-negative bacteria, which have a thinner peptidoglycan layer, while Gram-positive bacteria retain the stain due to their thicker cell wall. This step is critical for distinguishing between the two types of bacteria.