Microscopy in Class 11 Biology – Light & Electron Microscopes with Stains Explained

3.3 Microscopy

The discovery of cells and the study of their internal structures were only possible after the invention of the microscope.

Microscopy: The technique of using microscopes to observe and study objects too small to be seen with the naked eye. Microscopes use lenses and light or electron beams to magnify specimens.

Key Concepts in Microscopy

1. Magnification

Definition: The ability of a microscope to enlarge the image of an object.
Symbol: 'X', showing how many times the object is enlarged.
Example: 10X lens → enlarges a 1 µm object to 10 µm.
Total Magnification: Multiply the magnification of all lenses in the microscope.

2. Resolution

Definition: The ability of a microscope to distinguish between two points that are very close together.
Importance: Higher resolution → finer details can be observed.
Human Eye Resolution: ~0.1 mm
Light Microscope Resolution: ~250 nm (0.25 µm)
Electron Microscope Resolution: ~0.2 nm (0.0002 µm)

Types of Microscopes

A. Light Microscope

Uses light to illuminate the object.
Light passes through the specimen and two glass lenses:

Objective lens → produces enlarged image of object.
Ocular lens → further magnifies the image for viewing.

Magnification: Up to 1500X
Resolution: 0.2 µm
Advantages: Can view living organisms.
Limitations: Cells are often transparent, components not visible unless stained. Staining usually kills cells.

Major Parts of a Light Microscope:

Eyepiece (Ocular lens)
Objective lenses
Stage
Light source
Coarse & fine focus knobs
Mirror/illuminator

B. Electron Microscope (EM)

Uses electron beams instead of light.
Magnification & Resolution: Much higher than light microscope.
Limitation: Cannot view living cells (preparation kills specimen).

Types of Electron Microscopes:

Transmission Electron Microscope (TEM)

Passes electron beam through a thin specimen.
Used to view internal structures of cells.
Magnification: Up to 250,000X
Resolution: 0.2 nm

Scanning Electron Microscope (SEM)

Used to view surface details of cells or objects.
Surfaces are coated with metal, electron beam reflects off metal → image formed.
Magnification: Up to 100,000X
Resolution: 0.2 nm

Light Microscope vs Electron Microscope

Feature	Light Microscope	Electron Microscope
Illuminating source	Light (400–700 nm)	Electron beams (~1 nm)
Magnification	500X – 1500X	Direct: 16,000X; Photographic: 1,000,000X
Resolution	0.2 µm	0.2 nm
Specimens	Live & dead	Only dead & prepared specimens
Preparation time	Minutes – 1 hour	Several days
Image viewing	Through ocular lens	On zinc sulphate fluorescent screen

Stains in Microscopy and Their Role

Since many cell components are transparent, staining is used to enhance visibility of structures.

Common Stains and Uses:

Stain	Role / Target Structure
Methylene Blue	Stains nuclei and acidic components (DNA, RNA)
Iodine Solution	Stains starch granules and cytoplasm in plant cells
Eosin	Stains cytoplasm pink/red, helps contrast with nuclei
Hematoxylin	Stains nuclei blue/purple in tissue sections
Crystal Violet	Used in Gram staining to differentiate bacterial cell walls
Safranin	Counterstain in Gram staining, stains cell walls red/pink
Aceto-orcein / Acetocarmine	Stains chromosomes in mitosis and meiosis studies

Note:

Staining usually kills cells, except some special live-cell stains.
Stains help visualize organelles, nuclei, chromosomes, and cell structures under the microscope.

Additional Notes

Light microscopes → ideal for living cells, motility, and basic structure.
Electron microscopes → detailed ultrastructure, viruses, organelles, proteins.
Magnification ≠ Resolution: High magnification alone does not reveal fine details; resolution is crucial.

Key Points to Remember

Microscopy is essential for studying cell structure and function.
Light microscopes: live & simple; limited magnification & resolution.
Electron microscopes: ultra-high resolution, detailed; cannot study living cells.
TEM → internal structures; SEM → surface structures.
Stains enhance visibility of specific structures but mostly kill cells.

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