Histology Techniques and Staining Methods

Introduction

• The presentation is about histology, specifically methods of staining tissue samples for microscopic analysis.
• The presenter is Dr. Dalia Eita.
• The institution is Mansoura National University.

Agenda

• Methods for preparing tissue samples for light microscopy
• Staining principles using Hematoxylin & Eosin
• Types of stains
• Types of microscopes

Learning Objectives

• Demonstrate tissue fixation and embedding procedures.
• Identify and explain common staining techniques.
• Explain the application of cytochemical and histochemical approaches.
• Compare light and electron microscopy.

Special Stains for Organic Components

• Carbohydrates:
  • Periodic acid-Schiff (PAS) stain uses periodic acid to react with carbohydrates creating aldehyde groups, aldehyde group reacts with Schiff's reagent then turns magenta
  • Best Carmine stain highlights glycogen, giving glycogen granules a red colour.
• Lipids:
  • Frozen sections are used
  • Sudan III stains lipids orange
  • Sudan black stains lipids black
  • Osmium tetroxide stains lipids black

Other Methods of Staining

• Vital stains: These stains are used to observe living cells e.g. staining of phagocytes (cells of the body's immune system) with Trypan blue or Indian ink.
• Metachromatic Stains: These stains show differences in color compared to that of the dye. An example is mast cell's granules which have a reddish-purple color when stained with toluidine blue.
• Enzyme Histochemistry: Frozen tissue sections are used to locate and study specific enzymes e.g alkaline phosphatase in kidney tubules.

Quiz (PAS stain)

• PAS stains carbohydrates and turns magenta.

Microscopy: Types of Microscopes

• Light microscopes: Used in educational settings, have magnifications of 40x, 100x, 400x, and 1000x.
• Electron microscopes: Higher magnification (up to 50,000x) and resolution allowing for observation of smaller structures compared to light microscope.

Resolution Power

• Resolution power measures the smallest distance between two particles that can be distinguished as separate particles.
• Light microscopes have a resolution power of about 0.2 micrometers (200 nanometers).
• Electron microscopes have a resolution power of around 1 nanometer.

Practical Class: How to Use a Microscope

• Step-by-step instructions on how to use a light microscope, are provided in the form of instructions in the presentation.

Micro Technique for Electron Microscopy

• Ultrathin sections (50-100 nm thick) are made using an ultramicrotome.
• Sections are stained using heavy metals (lead citrate and uranyl acetate)
• Dark components in electron micrographs are called electron-dense, while light components are called electron-lucent.

Scanning Electron Microscope (SEM)

• Allows for 3-dimensional visualization of the surface of a fixed specimen

Explore the fascinating world of histology with this quiz on tissue staining techniques. Learn about the various methods for preparing tissue samples, principles of staining using Hematoxylin & Eosin, and the differences between light and electron microscopy. Ideal for students and professionals in the field of biological sciences.