Electron Microscopy

Electron Microscopy (EM)

Uses a beam of electrons instead of light to visualize specimens; provides much higher resolution than light microscopy.

Transmission Electron Microscope (TEM)

Developed in the 1930s; uses a very thin section of sample; electrons pass through the specimen to form an image → shows internal ultrastructure.

Scanning Electron Microscope (SEM)

Developed in the 1950s; electrons do not pass through but bounce off the sample surface → provides 3D surface imaging.

Light Microscope vs TEM vs SEM

LM uses light and glass lenses; TEM/SEM use electrons and electromagnetic lenses.

Fixation for TEM

Specimen fixed in glutaraldehyde and osmium tetroxide to preserve cellular structures.

Dehydration for TEM

Gradual alcohol dehydration: 50% → 70% → 80% → 90% → 95% → 100%.

Embedding for TEM

Specimen infiltrated and embedded in plastic resin before ultra-thin sectioning.

Sectioning for TEM

Sections cut using an ultramicrotome (~70 nm thick) and mounted on copper grids.

Plasmalemma (Cell Membrane)

Outer boundary of the cell; separates intracellular and extracellular fluids; composed of lipid bilayer with embedded proteins (Fluid Mosaic Model).

Fluid Mosaic Model

Describes the cell membrane as a flexible, moving bilayer of lipids with proteins floating within it.

Nucleus

Control center of the cell; contains genetic information; largest organelle; red blood cells lack nuclei; skeletal muscle cells are multinucleated.

Nuclear Envelope

Double membrane surrounding the nucleus; contains pores allowing molecular exchange.

Chromatin

Genetic material inside nucleus; heterochromatin (dense, inactive) and euchromatin (light, active).

Nucleolus

Site of ribosomal RNA (rRNA) synthesis inside the nucleus.

Endoplasmic Reticulum (ER)

Network of membranes continuous with the nuclear envelope; exists as rough (rER) and smooth (sER) types.

Rough Endoplasmic Reticulum (rER)

Has ribosomes; responsible for protein synthesis and transport.

Smooth Endoplasmic Reticulum (sER)

Lacks ribosomes; functions in lipid and steroid synthesis, detoxification.

Ribosomes

Small structures with large and small subunits; site of protein synthesis; can be free or attached to rER.

Golgi Apparatus

Series of flattened, membrane-bound sacs; modifies, packages, and sorts proteins for secretion or use in the cell.

Cis Face of Golgi

Receives vesicles from the ER (entry side).

Trans Face of Golgi

Shipping side; sends off modified proteins in vesicles.

Mitochondria

Powerhouse of the cell; double membrane with inner folds called cristae; site of ATP production.

Cristae

Inner membrane folds of mitochondria that increase surface area for ATP synthesis.

Mitochondrial DNA

Mitochondria contain their own DNA and reproduce independently by division.

Cytosol

Aqueous matrix of the cytoplasm excluding organelles; site of many chemical reactions like glycolysis and protein synthesis.

SEM Principle

Electron beam accelerated in a vacuum; interacts with metal-coated sample; scattered electrons detected to form a surface image.

SEM Components

Vacuum system, electron column, specimen chamber, and signal detection display with detectors and electronics.

SEM Specimen Preparation

Steps: fixation, dehydration (critical point drying), coating with metal.

SEM Magnification Range

1,000× to 100,000× magnification; produces 3D-like images.

Uses of SEM

Topography (surface texture), morphology (shape/size), composition (elemental makeup), and crystallography (atomic arrangement).

EDX (Energy Dispersive X-ray Analysis)

Technique used in SEM for elemental composition analysis of specimens via backscattered electrons.

Difference between TEM & SEM

TEM shows internal structure (thin section); SEM shows surface details (whole sample).

LM vs TEM vs SEM Images

LM uses visible light and color stains; TEM shows black-and-white internal ultrastructure; SEM provides 3D surface topography.

Example Specimens in EM

Red blood cells, mitochondria, Golgi apparatus, ER—each visible with different EM types.

Critical Point Drying

Method for SEM specimen dehydration to prevent collapse of surface structure.

Backscattered Electron Imaging

In SEM, distinguishes materials based on atomic number differences—higher atomic number = brighter area.