MR

Microscopy and Cell Division

Light Microscopy

  • Any kind of microscope that uses visible light to observe specimens.

The Compound Light Microscope

  • Ocular lens (eyepiece): remagnifies the image formed by the objective lens.

  • Body tube: transmits the image from the objective lens to the ocular lens.

  • Arm: supports the structure of the microscope.

  • Objective lenses: primary lenses that ‘ magnify the specimen.

  • Stage: holds the microscope slide in position.

  • Condenser: focuses light through the specimen.

  • Diaphragm: controls the amount of light entering the condenser.

  • Illuminator: light source.

  • Coarse focusing knob: used for large focus adjustments.

  • Fine focusing knob: used for precise focus.

  • Base: supports the microscope.

  • Stage adjuster (stage controls): moves the stage.

  • Rheostat: adjusts light intensity.

Path of Light and Image Orientation

  • Line of vision runs from the illuminator, through the condenser and specimen, through the objective lens, to the ocular lens.

  • What you see is flipped and upside down due to the optical pathway through the lenses.

  • The path of light (bottom to top) can be summarized as: Illuminator → Condenser → Specimen → Objective lenses → Ocular lens → Observer.

Compound Light Microscopy (Overview)

  • In a compound microscope, the image from the objective lens is magnified again by the ocular lens.

  • Total magnification is the product of the magnification powers of the objective lens and the ocular lens.

The Path of Light (Illustrative Layout)

  • Components include: ocular lens, body tube, objective lenses, specimen, condenser lenses, illuminator, base with source of illumination, prism.

  • The image orientation is flipped and upside down at the viewer due to the lens arrangement.

Resolution and Resolving Power

  • Resolution is the ability of the lenses to distinguish two points as separate.

  • A microscope with a resolving power of 0.4 nm can distinguish between two points at least 0.4 nm apart.

  • Expressed: \text{Resolution} = 0.4\ \text{nm}

Refractive Index and Oil Immersion

  • The refractive index is a measure of the light-bending ability of a medium.

  • Light may refract after passing through a specimen to an extent that it does not pass through the objective lens.

  • Immersion oil is used to keep light from refracting (reduces refraction between slide and objective).

  • Oil immersion objective lens requires immersion oil to maintain light path and improve resolution.

  • Without immersion oil, most light is refracted and lost; with immersion oil, unrefracted light is maximized.

Oil Immersion Setup (Key Points)

  • Oil is used with the oil immersion objective lens.

  • Immersion oil sits between the slide and the objective lens to minimize refraction.

  • The condenser, iris diaphragm, and light source work with the oil to maximize light collection.

Microscopes and Objective Lenses

  • Microscopes have several objective lenses:

    • 4X (scanning)

    • 10X (low power)

    • 40X (high power/high dry)

    • 100X (oil immersion)

  • The ocular lens (eyepiece) has a magnification of 10X.

  • Objective lenses have associated numerical apertures (e.g., 0.10, 0.25, 0.65, 1.25) and are paired with the ocular lens to determine total magnification.

Total Magnification and Examples

  • Total magnification is calculated by multiplying the magnification power of the objective lens by the magnification power of the ocular lens.

  • Examples (as given in the transcript):

    • 4X objective × 10X ocular = M_{total} = 4 \times 10 = 40X

    • 10X objective × 10X ocular = M_{total} = 10 \times 10 = 100X

    • 40X objective × 10X ocular = M_{total} = 40 \times 10 = 400X

Cell Diversity (The Cells as Basic Units)

  • There are over 250 different types of human cells.

  • Types differ in size, shape, and subcellular components; these differences lead to differences in functions.

  • Examples of cell types:

    • Fibroblasts

    • Erythrocytes

    • Skeletal muscle cell

    • Epithelial cells

    • Fat cell

    • Nerve cell

    • Macrophage

    • Sperm

    • Smooth muscle cells

  • (Classification by function):

    • (a) Cells that connect body parts, form linings, or transport gases

    • (b) Cells that move organs and body parts

    • (c) Cell that stores nutrients

    • (d) Cell that fights disease

    • (e) Cell that gathers information and controls body functions

    • (f) Cell of reproduction

Cells: The Smallest Living Units (3.1 Cells: The Smallest Living Units)

  • Cell diversity and specialization underlie organismal form and function.

  • This section introduces cell types and their general roles in physiology and tissue structure.

Meiosis and Reproduction (Garbled content from Page 14-15)

  • Meiosis is discussed in the transcript with references to:

    • Meiosis vs. mitosis; purpose to produce gametes.

    • Meiosis yields haploid gametes (not explicitly stated but implied by the context).

    • A tetrad is mentioned as part of meiosis terminology.

  • Embryo to birth development is referenced:

    • Stages listed include embryo, fetus, birth, infancy, toddler, childhood, youth, adulthood, old age (text appears garbled, but the developmental stages are indicated).

Interphase (1 of 5)

  • Interphase is the period from cell formation to cell division during which the cell carries out its routine activities and prepares for division.

  • During interphase, nuclear material is in an uncondensed chromatin state.

  • Interphase consists of subphases, which include the process of DNA replication (S phase).

The Cell Cycle (1–3) and Checkpoints

  • The cell cycle includes: G1 (Growth), S (DNA Synthesis), G2 (Growth and final preparations for division), and Mitosis (M) including Prophase, Metaphase, Anaphase, Telophase, followed by Cytokinesis.

  • G1 checkpoint (restriction point): ensures cells are ready for DNA synthesis.

  • G2/M checkpoint: ensures all DNA has been replicated and is not damaged before mitosis.

  • The diagrammatic flow: Interphase → G1 → S → G2 → Mitosis (Prophase, Metaphase, Anaphase, Telophase) → Cytokinesis.

The Cell Cycle (2 of 3) and Mitosis Overview

  • M phase is the mitotic phase during which division occurs; it consists of two events:

    • Mitosis (nuclear division)

    • Cytokinesis (cytoplasmic division)

  • The four stages of mitosis ensure each daughter cell receives a full copy of replicated DNA.

The Cell Cycle (Cell Division: Mitosis in Detail) (Pages 21–33)

  • Prophase: early and late phases

    • Early prophase: chromatin condenses into visible chromosomes; each chromosome has sister chromatids held at the centromere; centrosomes move to opposite poles; mitotic spindle forms; asters radiate from centrosome.

    • Late prophase: nuclear envelope breaks up; kinetochores attach to kinetochore microtubules and pull chromosomes toward the center; nonkinetochore microtubules push poles apart.

  • Metaphase: chromosomes align at the metaphase plate (midline of the cell); centromeres aligned at the equator.

  • Anaphase: centromeres split simultaneously; sister chromatids separate into individual chromosomes; kinetochore microtubules shorten and pull chromosomes toward opposite poles; nonkinetochore microtubules lengthen and push poles apart; chromosomes appear V-shaped as they move.

  • Telophase: chromosomes arrive at poles; chromatin decondenses; new nuclear envelopes form around each chromatin mass; nucleoli reappear; spindle breaks down.

  • Cytokinesis: cytokinesis begins in late anaphase and continues through mitosis; a contractile ring of actin forms a cleavage furrow that pinches the cell into two daughter cells.

Mitosis: Focus Figures and Phases (Additional Details from Figures)

  • Primary features depicted in FOCUS FIGURE 3.4 Mitosis include:

    • Centrosomes with two centrioles and the spindle apparatus.

    • Chromosomes consisting of sister chromatids held at the centromere.

    • Kinetochore microtubules interacting with kinetochores.

    • Asters and mitotic spindle dynamics guiding chromosome movement.

  • The figure sequence shows the transformation from Interphase through Prophase, Metaphase, Anaphase, Telophase, and Cytokinesis with corresponding structural changes.

Interphase vs. Mitosis: Summary (Unnumbered Figure 3.10 Page 100)

  • Interphase is the period when the cell carries out normal metabolic activities and grows; it is not part of mitosis.

  • During interphase, the DNA-containing material is in the form of chromatin; the nuclear envelope and nucleoli are intact.

  • The three distinct periods of interphase are G1, S, and G2.

  • Prophase and the subsequent stages are part of mitosis, culminating in cytokinesis.

  • The micrographs illustrate dividing lung cells from a newt with chromatin in blue and microtubules in green; red fibers are intermediate filaments.

Mitosis in Animal Cells: Whitefish Blastula (Figure References)

  • The transcript references Mitosis in animal cells illustrated via Whitefish blastula at various magnifications (e.g., 400x, 600x, 1000x) across Interphase, Prophase, Metaphase, Anaphase, Telophase.

  • These images illustrate the progression of chromosomal condensation, alignment, separation, and cytokinesis in a rapid developmental model.

Practical and Real-World Relevance

  • Light microscopy is fundamental to observing cell structure, tissue organization, and basic cellular processes in biology labs.

  • Understanding total magnification and resolution is essential for selecting appropriate objectives and interpreting microscopic images.

  • The cell cycle and mitosis are core concepts in growth, development, and tissue repair, with dysregulation implicated in cancer and developmental disorders.

  • Oil immersion techniques demonstrate how physics of light and refractive indices influence image clarity and resolution in microscopy.

Summary of Key Equations and Numbers

  • Total Magnification: M{total} = M{objective} \times M_{ocular}

    • Examples:

    • 4X objective with 10X ocular: M_{total} = 4 \times 10 = 40X

    • 10X objective with 10X ocular: M_{total} = 10 \times 10 = 100X

    • 40X objective with 10X ocular: M_{total} = 40 \times 10 = 400X

  • Resolution: \text{Resolution} = 0.4\ \text{nm}

  • Common objective magnifications: 4X (scanning), 10X (low power), 40X (high power/high dry), 100X (oil immersion)

  • Ocular magnification: 10X

  • Refractive index concepts and oil immersion techniques help maintain image clarity by reducing light refraction.