Microscopy: Structure, Use, and Principles for Anatomy & Physiology (lab)

Microscope Structure and Function

Introduction

The compound light microscope is an essential tool in Anatomy & Physiology for observing cells and tissues at high magnification. Understanding its parts, handling, and principles of operation is crucial for accurate and safe laboratory work.

Parts of the Compound Microscope

• Ocular lenses (Eyepieces): The lenses you look through, typically with 10x magnification.

• Objective lenses: Mounted on a rotating nosepiece, usually offering 4x, 10x, 40x, and sometimes 100x magnification.

• Arm: The supporting structure connecting the base and the head; used for carrying the microscope.

• Base: The bottom support of the microscope.

• Mechanical stage: Platform where slides are placed; includes stage clips and controls for precise movement.

• Condenser: Focuses light onto the specimen.

• Diaphragm lever: Adjusts the amount of light passing through the specimen.

• Coarse adjustment knob: Moves the stage up and down for general focusing (use only with lowest power objective).

• Fine adjustment knob: Allows for precise focusing, especially at higher magnifications.

• Substage light: Provides illumination from below the stage.

• Light control: Adjusts the intensity of the substage light.

Proper Handling and Care

• Carry the microscope upright, supporting it with one hand on the arm and the other under the base.

• Do not swing or jar the instrument when transporting or setting it down.

• Clean lenses only with lens paper, using a gentle circular motion.

• Clean lenses before and after use.

• Always begin focusing with the lowest power objective lens.

• Use the coarse adjustment knob only with the lowest power objective lens.

• Use a cover slip with wet mounts.

• Remove slides from the stage, rotate to the lowest power objective, return oculars to the 'stow position,' wrap the cord, and replace the dust cover after use.

• Never remove any parts from the microscope.

• Report any problems to the instructor immediately.

Principles of Microscopy

Magnification and Resolution

Microscopes allow for the visualization of small structures by magnifying images and resolving fine details.

• Magnification: The process of enlarging the appearance of an object. Total magnification is calculated by multiplying the magnification of the ocular lens by that of the objective lens. Formula: $\text{Total Magnification} = \text{Ocular Magnification} \times \text{Objective Magnification}$

• Resolution: The ability to distinguish two close points as separate entities. Higher resolution allows for clearer, more detailed images.

Field of View and Working Distance

• Field of View: The visible area seen through the microscope. As magnification increases, the field of view decreases.

• Working Distance: The distance between the objective lens and the specimen. Higher magnification lenses have shorter working distances.

Depth of Field

• Depth of Field: The thickness of the specimen that is in focus at one time. Lower magnification provides greater depth of field; higher magnification reduces it.

Parfocality

• Parfocal: A feature of microscopes that allows specimens to remain in focus when switching between objective lenses with minimal adjustment.

Measuring Field Diameter and Estimating Object Size

Field Diameter Measurement

• Field diameter can be measured for one objective lens and calculated for others using the following formula:

Formula: $\text{Field Diameter}_{A} \times \text{Total Magnification}_{A} = \text{Field Diameter}_{B} \times \text{Total Magnification}_{B}$

• Knowing the field diameter allows estimation of the size of objects in the field. For example, if the field diameter is 4 mm and an object spans half the field, its estimated size is 2 mm.

• Specimens are usually measured in micrometers (μm) and millimeters (mm), which are part of the metric system.

Summary Table: Microscope Parts and Functions

Part	Function
Ocular lens	Magnifies image, usually 10x
Objective lenses	Provide various magnifications (4x, 10x, 40x, 100x)
Arm	Support and carrying handle
Base	Stability and support
Mechanical stage	Holds and moves slide
Condenser	Focuses light on specimen
Diaphragm lever	Controls light intensity
Coarse adjustment knob	General focusing (low power only)
Fine adjustment knob	Precise focusing
Substage light	Illuminates specimen
Light control	Adjusts light brightness

Key Relationships in Microscopy

• Increasing magnification decreases field of view and working distance.

• Depth of field is greater at lower magnifications.

• Proper focusing technique and care are essential for accurate observation and instrument longevity.

Example Application

When observing a cheek cell at 40x objective (total magnification 400x), the field of view is much smaller than at 4x objective (total magnification 40x). Accurate measurement and focusing are necessary to study cell structure.

Additional info: These principles are foundational for all microscopy work in Anatomy & Physiology, including histology and cytology labs.