Generalities of Histology and Fundamental Tissues Study Guide

Academic Agenda for Unit 2: Developmental Biology

The academic activities for Unit 2 are divided into two groups, following a specific schedule throughout the months of August and September 2026. For Group 1, the schedule begins on August 11 with the class on Histology Generalities and Fundamental Tissues. This is followed on August 18 by the class on Epithelial Tissue and a quiz on histology generalities. On August 25, the group covers Connective Tissue and Cartilage along with an epithelial tissue quiz. The laboratory session for fundamental tissues is held on August 28. Review activities for Unit 2 take place on September 1, and the final unit exam is scheduled for September 8.

For Group 2, the schedule starts on August 13 with Histology Generalities and Fundamental Tissues. The Epithelial Tissue class and histology generalities quiz occur on August 20. On August 27, the group studies Connective Tissue and Cartilage and takes the epithelial tissue quiz. Similar to Group 1, the laboratory for fundamental tissues occurs on August 28, the unit review activity on September 1, and the Unit 2 partial exam on September 8. All sessions are led by Professor Liliana Sofía Martínez Acevedo from CES University.

Fundamental Concepts of Histology and the Cell

The primary objective of histology is to understand the microanatomy of cells, tissues, and organs, specifically correlating their physical structure with their biological function. Histology, derived from the Greek work "histos" meaning tissues and "logia" meaning science, is also known as microscopic anatomy. It is defined as the scientific study of the microscopic structures of the tissues and organs of the body.

The cell is the basic structural and functional unit of all multicellular organisms. These cells do not exist in isolation but organize themselves into tissues. Tissues, in turn, form organs, which then organize into apparatuses or systems to constitute a multicellular living being. Cells within these structures join together to carry out a common, specialized function.

Histotechnology and the Acquisition of Biological Samples

Histotechnology involves the preparation of materials from biological tissues to be visualized as histological preparations or slides. This process allows for detailed histological description. However, several critical aspects must be considered when obtaining a tissue for study. These include the state of the animal (whether it is alive or a cadaver), the species (noting characteristics like hair or fat layers), and the method of acquisition, which requires minimal manipulation of the tissue. The time of the sample collection should not exceed 24 hours, and conservation temperatures must be kept low. Proper preservation is essential to avoid autolysis, which can be achieved through refrigeration, freezing, or using a $10\%$ formalin solution.

Autolysis, coming from the Greek "autos" (self) and "lysis" (dissolution), is an anaerobic biological process where a cell destroys itself. This is a spontaneous cell lysis process typically caused by the activity of lytic proteins known as autolysins. Effective histotechnology relies on stopping this process immediately to preserve the structural integrity of the sample.

The Ten Steps of the Histotechnology Process

The first step is fixation, which aims to maintain the histological structure of the tissue. The standard agent used is $10\%$ buffered formalin, which preserves cellular substances and structures, stops autolysis, hardens the tissue, and destroys pathogenic microorganisms. For effective fixation, the tissue sample should have a maximum size of $1\,cm^3$, following a ratio of 1 part tissue to 9 parts formalin, for a duration of 24 hours.

The second step is manual cutting, where the tissue is sectioned into smaller pieces that fit into a histological cassette, also known as a histocassette. Third, the sample undergoes dehydration and clearing. Dehydration is achieved by passing the histocassette through progressive immersions in gradual alcoholic solutions of increasing concentration (Ethyl alcohol at $60\%$, $70\%$, $80\%$, $96\%$, and $100\%$) to avoid sudden dehydration. Clearing involves the extraction of alcohol using organic and aromatic solvents called clearing agents, such as xylol, toluene, benzene, or chloroform.

The fourth step is pre-inclusion in paraffin, using paraffin as a liquid organic compound that serves as a support upon solidification. Fifth is inclusion in paraffin, where a paraffin dispenser equipment maintains the substance in a liquid state at a temperature between $56$ and $58\,^{\circ}C$. A cooling plate set at $0\,^{\circ}C \pm 1\,^{\circ}C$ is then used to solidify the material into paraffin blocks containing the tissues.

Sixth, microtomy is performed to obtain thin sections using a microtome. These cuts are typically between $3$ and $10\,\mu m$ thick, where $1\,\mu m$ is the thousandth part of a millimeter. Seventh, the sections undergo flotation and adhesion. They are extended and floated in a hot water bath at $40$ to $45\,^{\circ}C$ containing gelatin or another adhesive agent before being mounted onto a glass slide.

Eighth is deparaffinization. The slides are leaned obliquely to drain excess liquid and then placed horizontally in an oven at $50\,^{\circ}C$ for 2 to 4 hours. This evaporates the liquid and allows the adhesive substance to firmly bond the tissue cut to the slide. Ninth is staining and coloration, a process that makes cells visible by applying dyes or chemical agents. Hematoxylin and eosin (H&E) is the most common method. Finally, the tenth step is mounting, where the tissue section is covered with a drop of adhesive resin and a glass coverslip.

Chemical Foundations of Staining and Microscopy

Staining relies on the chemical affinity between cellular components and dyes. Acidic components of a cell, such as nuclear chromatin or chromosomes, have an affinity for basic dyes like Hematoxylin. These components are described as basophilic or hematoxylinophilic and appear blue-purple. Conversely, basic components of a cell, such as the cytoplasm and intercellular substance, take up acidic dyes like Eosin. These components are described as acidophilic or eosinophilic and appear pink.

Microscopy is the use of an optical microscope, an instrument that amplifies images to permit the observation of details not visible to the naked eye. Histological observation requires a mental reconstruction of structures from a bidimensional view back into their original tridimensional organization through various planes of cut. The optical microscope consists of several parts: the base (support); the arm (holds optical parts and focus mechanisms); the stage (where the slide is placed); the ocular tube or binocular head (holding the ocular lens); the nosepiece or revolver (rotating piece for objectives); and the condenser screw (adjusts condenser position).

Additional microscope components include the macrometric and micrometric screws located on the arm to move the stage toward or away from the objective lens rapidly or slowly. The ocular lens increases the image produced by the objectives, which are cylinders containing lenses with magnifications of $4X$, $10X$, $40X$, and $100X$. The condenser illuminates the preparation, while the diaphragm controls the amount of light passing through. The light source, located at the base or below the condenser, provides the required illumination.

The Four Fundamental Tissue Types

All organs in the body are composed of four basic types of tissues. Epithelial tissue covers body surfaces, lines body cavities, and forms glands. It is classified based on the number of cell layers and the shape of the cells, often possessing specific surface modifications.

Connective tissue underlies or structurally and functionally supports the other three basic tissues. It can specialize into structures such as bone and cartilage. Muscular tissue is composed of contractile cells and is responsible for movement. It is differentiated into three types: smooth muscle, striated (skeletal) muscle, and cardiac muscle. Finally, nervous tissue receives, transmits, and integrates information from both the internal and external environments to control body activities, primarily through specialized nerve cells called neurons.