Topic 2: Introduction to Cytology and Methods of Cell Study; Principles of cytological and histological investigation, cytochemistry, immunohistochemistry and in situ hybridization.
Covers the principles of cytological and histological investigation and how dyes bind to cell components through acidic and basic staining chemistry. It then goes through staining methods for carbohydrates (the PAS reaction), lipids and proteins, followed by immunohistochemistry with direct and indirect antibody methods, in situ hybridization, and how the choice of microscope depends on the technique used.
Topic 2 of 59 in the MU Pleven syllabus for Cytology (Medicine, Year 1). The free AceMed note covers it in 10 sections, about 340 words (3 min of reading), with 28 flashcards.
In this note
Topic 2: Introduction to Cytology and Methods of Cell Study
Principles of Cytological and Histological Investigation
Histochemistry: Definition and Staining Principles
The Chemical Basis of Staining
Demonstrating Carbohydrates:
Demonstrating Lipids
Demonstrating Proteins
Immunohistochemistry (IHC)
In Situ Hybridization (ISH)
Connecting Techniques to Microscopy
Principles of Cytological and Histological Investigation
Cytology and histology study the structure and chemical composition of cells and tissues.
Microscopy is the core tool for visualization. The type of microscope (e.g., light, fluorescence) is chosen based on the technique and the required detail.
Histochemistry: Definition and Staining Principles
Histochemistry blends chemistry and biology to visually study the location of specific molecules in tissues.
It relies on staining techniques where dyes selectively bind to target molecules.
The Chemical Basis of Staining
Acidic Dyes (e.g., Eosin):
Carry a net negative charge.
Bind to cationic groups (e.g., proteins), a property called acidophilia.
Stains cytoplasmic filaments, intracellular membranes, and extracellular fibers.
Basic Dyes (e.g., Hematoxylin):
Carry a net positive charge.
Bind to anionic groups (e.g., nucleic acids), a property called basophilia.
Stains the cell nucleus, rough ER, and cartilage matrix.
Demonstrating Carbohydrates:
The PAS Reaction
Carbohydrates: Biomolecules like glycogen and cellulose.
Periodic Acid-Schiff (PAS) Reaction:
Mechanism: Oxidation converts hydroxyl groups to aldehydes. Schiff's reagent reacts with them, producing a purple-magenta colour.
Applications: Visualizing goblet cells (mucin), glycogen in liver/muscle, and the cell surface glycocalyx.
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