Staining is a technique used to identify target molecules and provide contrast in an image, usually in microscopy. For example, biological tissues ranging from muscle fibers to organelles, are often stained to highlight particular regions for viewing with a microscope. Overall, staining involves adding a class specific dye, such as DNA, proteins, lipids or carbohydrates, to quantify and qualify their presence.
There are two common types of staining in biochemistry:
In vivo staining involves dyeing living tissue by causing cells or structures to take up the stain’s color. In doing so reveals
In vitro staining is done outside of the biological context. In vitro staining is usually used with other methods such as fixation and sample preparation. For example, the crystal violet stain only works with Gram-positive bacteria. If then, another stain is used that works on all cell-types, then Gram-negative bacteria can be identified.
In Vitro Methods
In vitro methods vary given the biological materials at hand, but in general, they follow these steps:
Preparation, either through
Fixation: preserve the shape of cells or tissues
Permeabilization: applying a mild surfactant to dissolve cell membranes, allowing for greater access to organelles
Mounting: applying a sample to a glass microscope slide.
Immerse target sample in the stain solution, and rinsing to remove excess dye.
If this is ineffective, then a mordant is usually required; a chemical compound which reacts with the stain to produce a colored precipitate.
Some common examples of staining techniques are:
Gram Staining: Determines gram status to classify bacteria, which are either Gram-negative or Gram-positive. Based on cell wall composition, Gram-positive bacteria stain a dark blue to violet, whereas Gram-negative appear red or pink.
Haematoxylin and eosin staining: Used in histology to examine thin sections of tissues. Hematoxylin targets cell nuclei and stains them blue, whereas eosin stains the cytoplasm pink.
Masson’s Trichrome: a three-color staining method used to distinguish cells from connective tissue.
Types of Stains:
Acridine Orange: a fluorescent cationic dye for cell cycle determination targeting nucleic acids
Coomassie blue: a stain used in gel electrophoresis that stains proteins blue
Crystal Violet: stains cell walls purple, and is the primary stain used in Gram staining
DAPI: binds to DNA, and shines a blue fluorescence when viewed under ultraviolet light
Eosin: a counterstain to hematoxylin and to color cytoplasmic material
Ethidium Bromide: colors DNA a fluorescent red-orange color.
Hematoxylin: targets the nucleus, and is the counter part to eosin
Iodine: indicates the presence of starch
Methylene blue: stains the nucleus of animal cells