Microscopy: Principles, Structure, and Use in Anatomy & Physiology

Lab I: The Microscope

Objectives

This section outlines the key learning goals for mastering the compound microscope in anatomy and physiology laboratory settings.

• Identify the parts of the compound microscope and state the function of each.

• Determine the magnifying power of a microscope given the powers of the ocular and objective lenses.

• Learn how to use the microscope properly for examination of slides and troubleshooting common difficulties.

• Describe the relationship between magnification and field of view, and between magnification and depth of field.

• Make a temporary wet mount slide.

• Know the purpose of staining specimens.

• Know when and how to use the microscope’s oil immersion lens properly.

• Know how to clean and care for a microscope.

Introduction to the Microscope

Overview

The compound microscope is an essential tool for observing specimens that are too small for the unaided eye. Effective use of the microscope enhances laboratory experience and is foundational for biology and anatomy studies. Understanding its structure and function is crucial for future coursework and careers in science.

Background: Light and Lenses

Microscopes utilize the principles of light refraction and lens curvature to magnify images. Light travels in straight lines but bends when passing from one medium to another (e.g., water to air), a phenomenon called refraction. Curved glass surfaces, known as lenses, bend light to focus and magnify images.

• Refraction: The bending of light as it passes between different media (see Figure 1).

• Lens: A curved glass surface that focuses light to produce a magnified image.

The Purpose of the Microscope

Key Concepts

Microscopy relies on three main principles: magnification, resolution, and contrast.

• Magnification: The degree to which the image of a specimen is enlarged. Total magnification is calculated by multiplying the magnification of the objective lens by that of the ocular lens.

$\text{Total Magnification} = \text{Objective Magnification} \times \text{Ocular Magnification}$

• Resolution: The ability to distinguish two points as separate entities. Higher resolution allows for greater detail in the image.

• Use of Oil: Oil immersion is used with high-power objectives (typically 100X) to increase resolution by reducing light refraction between the slide and the lens.

• Contrast: The ability to see specimen detail against its background. Staining specimens or adjusting the iris diaphragm can improve contrast.

Parts of the Compound Microscope

Support Structures and Carrying the Microscope

The base is the lowest part, providing stability. The arm connects the base to the head and is used for carrying the microscope.

• Base: Provides stability and support.

• Arm: Used for carrying; connects base to head.

• Stage: Platform where slides are placed; equipped with stage clips and XY controls for movement.

Focus Adjustments

• Coarse Focus Knob: Used for large adjustments, especially under low power.

• Fine Focus Knob: Used for precise, small adjustments, especially under high power.

Lighting and Adjusting the Light

• Light Source: Provides illumination for the specimen.

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

• Condenser Lens: Focuses light onto the specimen.

Lenses and Optics

• Objective Lenses: Attached to the revolving nosepiece; typically 4X, 10X, 40X, and 100X (oil immersion).

• Ocular (Eyepiece) Lens: Usually 10X magnification; produces a virtual image.

When using the microscope, the final image is formed by the lens of your eye, which converts the virtual image to a real image on the retina.

Table: Main Parts of the Compound Microscope and Their Functions

Part	Function
Base	Supports the microscope
Arm	Used for carrying; connects base to head
Stage	Platform for slides; allows movement in XY direction
Coarse Focus Knob	Large adjustments to focus
Fine Focus Knob	Small, precise adjustments to focus
Light Source	Illuminates specimen
Iris Diaphragm	Regulates light intensity
Condenser Lens	Focuses light onto specimen
Objective Lenses	Magnify specimen (4X, 10X, 40X, 100X)
Ocular Lens	Further magnifies image (usually 10X)
Revolving Nosepiece	Holds and rotates objective lenses

Using the Microscope

Basic Steps

Proper use of the microscope involves several key steps:

1. Carry the microscope with both hands (one on the arm, one under the base).

2. Place the slide on the stage and secure with stage clips.

3. Start with the lowest power objective lens (usually 4X or 10X).

4. Use the coarse focus knob to bring the specimen into view, then use the fine focus knob for clarity.

5. Adjust the iris diaphragm and condenser for optimal lighting and contrast.

6. Switch to higher power objectives as needed, refocusing with the fine focus knob.

7. For oil immersion (100X), place a drop of oil on the slide before rotating the lens into position.

Care and Maintenance

• Clean lenses with lens paper only.

• Store the microscope with the lowest power objective in place.

• Cover the microscope when not in use to prevent dust accumulation.

Additional info:

• Staining specimens increases contrast and allows for better visualization of cellular structures.

• Field of view decreases as magnification increases; depth of field also decreases, requiring careful focusing.

• Microscope use is foundational for many laboratory techniques in anatomy and physiology.