Enhances contrast in unstained, transparent specimens by converting phase shifts (due to variations in refractive index and thickness within the specimen) into amplitude (brightness) differences.

It improves the visibility of internal structures without staining but does not inherently increase the resolving power beyond the limits of bright-field microscopy. Its primary role is to improve contrast, making structures distinguishable that would otherwise be invisible.

4.2 Phase Contrast Microscope

A type of light microscope that uses phase differences in light to create contrast in transparent specimens.

Principle: Light passing through a specimen undergoes phase shifts (retardation) proportional to the refractive index and thickness of the sample. The phase contrast microscope converts these invisible phase differences into visible differences in brightness.
Components:
- Annular Diaphragm: Located in the condenser, it produces a hollow cone of light.
- Phase Plate: Located in the objective lens, it has a phase ring that retards or advances the phase of the direct (unscattered) light, while diffracted (scattered) light passes through unaltered.
Working: Unscattered light passes through the phase ring, undergoing a phase shift. Scattered light bypasses the phase ring. When these two sets of waves recombine, their interference patterns create variations in amplitude (brightness), making transparent structures visible as darker or brighter areas against a background.
Applications: Observing living, unstained cells, cell division, and intracellular structures.

4.3 Fluorescent Microscopy

A type of light microscopy that uses fluorescence instead of, or in addition to, reflection and absorption to study properties of organic or inorganic substances.

Principle: Specimens are stained with fluorophores (fluorescent dyes) that absorb light at a specific wavelength (excitation wavelength) and then emit light at a longer, lower-energy wavelength (emission wavelength).
Working:
1. An excitation light source (e.g., mercury lamp, LED, laser) emits light of the appropriate wavelength.
2. An excitation filter selects only the excitation wavelength to illuminate the specimen.
3. The fluorophores in the specimen absorb this light and emit fluorescent light.
4. A dichroic mirror reflects the excitation light towards the specimen and allows the longer-wavelength emitted fluorescent light to pass through to the eyepiece/detector.
5. An emission filter blocks any remaining excitation light and passes only the emitted fluorescent light, creating a bright image of the fluorescent structures against a dark background.
Diagram (Conceptual):