Microbiology
Gram Characteristics of Bacteria Introduction Bacteria are interesting little critters in that they are not easily visualized without some sort of stain. In classical methodology, two different kinds of stains have been used to identify bacteria: acidic and basic dyes. Acidic dyes are so called because they consist of a salt that has a cation that transfers no color, but the anion portion is colored and does give off color. Basic dyes consist of an anion that does not give off any color and a cation that does stain biological samples. Examples of acidic dyes include acid fuchsin and eosin; examples of basic dyes include crystal violet, methylene blue and safranin. As a general rule, basic dyes are attracted to the surface of a bacterial cell (due to ionization of the carboxyl groups in the fatty and amino acids present in the cell wall/membrane). Basic dyes are attracted to the nucleic acid component of the bacterial cell, as well. As a general rule, acidic dyes are attracted to basic (alkaline) subcellular components. The most common stains used to stain bacteria are basic dyes. The most common background stains are acidic dyes since they do not cross the cell wall/membrane particularly easily. A background stain is a stain used to stain everything other than the bacteria. The Gram stain is one of the oldest, most cost-efficient, yet most under-utilized, staining method used to identify bacteria. It consists of a basic dye (the primary stain crystal violet), a mordant (Gram 's iodine), decolorizer (acetone:alcohol) and a counter stain (safranin). A mordant is a compound which helps hold the primary stain to the bacteria. In this case, the iodine forms a complex with the crystal violet to "lock it into" the cell wall/membrane. The decolorizer is used to decolorize the bacteria that "do not like" the primary stain. The counter stain is used to stain those bacteria which were decolorized by the acetone:alcohol. Bacteria which retain the crystal violet:iodine complex
References: 1. Beishir, L.: Microbiology in Practice: A Self-Instructional Laboratory Course, Fifth Edition. (Harper Collins: New York) 1991. 2. Jawetz, Melnick and Adelberg: Medical Microbiology, Nineteenth Edition. (Appleton and Lange: Norwalk, CT) 1991. 3. Tortora, Case and Funke: Microbiology: An Introduction, Fourth Edition. (Benjamin Cummings: Redwood City, CA) 1992. 4. Zubay: Biochemistry. (Addison Wesley: Reading, MA) 1983. 4