Which of the following answers lists the steps of gram-staining in the correct order?

• A. Stain with primary stain (crystal violet), add iodine, add decolorizer, stain with counterstain (safranin).
• B. Add iodine, add decolorizer, stain with primary stain (crystal violet), stain with counterstain (safranin).
• C. Stain with primary stain (crystal violet), add decolorizer, stain with counterstain (safranin), add iodine.
• D. None of the above are correct.

Answer: (A)

Verified step by step guidance

1. Understand the purpose of Gram staining: It is a differential staining technique used to classify bacteria into two groups: Gram-positive and Gram-negative, based on the characteristics of their cell walls.
2. Identify the primary stain: The process begins with the application of crystal violet, which is the primary stain that colors all the cells.
3. Apply the mordant: Iodine is added after the primary stain. It acts as a mordant, forming a complex with crystal violet that helps the dye adhere to the cell walls.
4. Use the decolorizer: An alcohol or acetone solution is used to decolorize the cells. This step differentiates the cells; Gram-positive cells retain the crystal violet-iodine complex, while Gram-negative cells lose the color.
5. Apply the counterstain: Finally, safranin is used as a counterstain. It stains the decolorized Gram-negative cells pink, while Gram-positive cells remain purple.