**2.1 Classical dyes**

According to the definition given by the Merriam-Webster dictionary, a dye or a stain is able to penetrate living cells or tissues without inducing immediate obvious degenerative changes and thus, it is also called a vital stain. Supravital staining implies the removal of living cells from an organism, whereas intravital staining involves injecting (or otherwise administering) the dye into the organism. The term "vital stain" is sometimes used to refer to an intravital stain, and in some other situations, it is interchangeable with a supravital stain, the main idea being that the cell being looked at is still alive. In a more strict way of speaking, the term "vital staining" has a meaning which is opposite to "supravital staining." If living cells take up the stain during supravital staining, living cells exclude the dye during "vital staining"; for example, they color negatively and only dead cells color positively, and hence, viability can be measured by counting in percentage the amount of total cells that stain negatively. Because the nature of the dye defines if the staining is either supravital or intravital, a mix of supravital and vital dyes can be employed to better categorize cells into various groups (e.g., viable, dead, dying) (**Figures 1** and **2**).

**Figure 1.** *Metallic green sheen characteristic colonies of Escherichia coli on eosin methylene blue agar (EMB) in close-up.*

**Figure 2.** *A vial of methylene blue.*

*The Use of Indocyanine Green in Colorectal Surgery DOI: http://dx.doi.org/10.5772/intechopen.100301*

Tissue staining, also known as chromoscopy, can be used as an adjuvant technique in gastrointestinal endoscopy to help with the recognition of subtle lesions, such as is the case with polyps or, more so, allows to directly target biopsies, which may happen in the case of Barrett's esophagus, in order to increase the precision of the diagnosis. Four endoscopic staining techniques have been described—vital staining (the use of an agent that is absorbed by the intestinal epithelium), contrast staining (the use of a substance to accentuate the aspect of the surface), reactive staining (the use of an agent that can fire chemical reactions), and tattooing (a technique using agents such as India ink to underline a special lesion on the mucosa).

Tissue stains used in gastrointestinal surgery, as mentioned by Fennerty [1], can be classified as follows:

a.tattooing agents


b.absorptive stains

	- 1.indigo carmine and cresyl violet (accentuates mucosal topography, allowing recognition of abnormal small bowel sprue and colonic mucosa inflammatory bowel disease, polyps).
	- 1.Congo red (identifies acid-secreting portions of the stomach postoperatively and documents achlorhydria) and
	- 2.phenol red (identifies alkaline areas of the stomach).

#### **2.2 Fluorescent dyes**

According to the definition offered by www.britannica.com, fluorescence is the emission of electromagnetic radiation, usually visible light, caused by the excitation of atoms in a material, which then reemit almost immediately (within about 10 s). The initial excitation is frequently determined by the absorption of energy from

#### **Figure 3.**

*High-resolution fluorescent microscope image of clusters of tumor cells in red surrounded by normal cells and normal skin in green. Photo source: www.shutterstock.com.*

#### **Figure 4.**

*Research sample slide of tumor tissue where cells have been stained for different proteins using a fluorescently tagged antibody. Presented as false-color image on white background.*

incident radiation of particles, as is the case with X-rays or electrons (**Figure 3**). Due to the fact that reemission happens so quickly, the fluorescence stops when the exciting source is removed, unlike the phenomenon of phosphorescence (**Figure 4**), which later persists as an afterglow.
