Microscopy

Major Parts of the Microscope

• Eyepiece: The lens you look through, typically magnifying 10X. It is crucial for viewing the specimen clearly.

• Objectives: Different lenses with varying magnifications (4X, 10X, 40X) that allow for detailed examination of specimens at different levels of detail.

• Coarse Focus Knob: Adjusts the stage height significantly for initial focusing on the specimen.

• Fine Focus Knob: Provides precise adjustments for clear focusing, essential for high magnification.

• Light Source: Illuminates the specimen, critical for visibility and clarity.

• Stage and Stage Clips: The flat platform where slides are placed, with clips to hold the slides securely.

Types of Microscopes

• Light Microscope: Magnifies up to 200X, uses visible light and lenses to magnify specimens. Ideal for viewing larger cells and tissues.

• Electron Microscope: Uses electron beams for magnification over 1 million X, allowing for visualization of smaller structures like organelles. Requires complex sample preparation and is expensive.

• Comparison: Light microscopes are more accessible and easier to use, while electron microscopes provide much higher resolution and detail.

Setting Up and Using the Microscope

• Slide Preparation: Properly prepare slides by placing specimens on a glass slide and covering them with a cover slip to avoid air bubbles.

• Focusing: Start with the lowest power objective and use the coarse focus knob to locate the specimen, then switch to higher magnifications using the fine focus knob.

• Light Adjustments: Adjust the diaphragm to control the amount of light passing through the specimen for optimal visibility.

• Wet mount: Clean a microscope slide. Add a few drops of liquid mixed with bacteria to the slide. Place a cover slip over the sample on the slide. Use a drop of water to suspend the specimen between the slide and cover slip. Lower the cover slip into place using a toothpick or equivalent.

Cell Biology

Cell Definition and Features

• Definition: A cell is the smallest living unit, discovered by Robert Hooke in 1665. Cells are the building blocks of all organisms.

• Cytology: The study of cellular structure and function, essential for understanding life processes.

• Cell Theory: States that all living organisms are composed of cells, cells arise from pre-existing cells, and cells are the basic unit of life.

Prokaryotic vs. Eukaryotic Cells

• Prokaryotic Cells: have a non-membrane-bound nucleus, smaller (1 micrometer), and include bacteria. They have a nucleoid region for DNA.

• Eukaryotic Cells: Have a membrane-bound nucleus, larger (10-100 micrometers), and include plants and animals. They contain organelles.

• Key Differences: Prokaryotes are generally unicellular, while eukaryotes can be unicellular or multicellular. Eukaryotes have membrane-bound organelles, unlike prokaryotes.

Hierarchical Organization of Life

• Levels of Organization: Ranges from atoms to biosphere, illustrating the complexity of biological systems: Atom < Molecule < Organelle < Cell < Tissue < Organ < Organ System < Organism < Population < Community < Ecosystem < Biosphere.

• Significance: Each level builds on the previous one, demonstrating the interdependence of biological structures and functions.

Organelles

Structure and Function of Organelles

• Cell Wall: Found in plant cells, fungi, and bacteria; provides structure and protection. It is rigid and non-living.

• Cell Membrane: Present in all cells; regulates entry and exit of substances, maintaining homeostasis.

• Nucleus: The control center of eukaryotic cells, housing DNA and regulating cell activities.

• Mitochondria: The powerhouse of the cell, generating ATP through cellular respiration.

• Chloroplasts: Found in plant cells, responsible for photosynthesis and energy production.

• Nucleolus: spherical body within the nucleus of most eukaryotic cells, involved in the synthesis of ribosomal RNA (rRNA) and the formation of ribosomes.

• Ribosomes: Present in all cells, essential for protein synthesis.

• Cytoplasm – found in all cells. Functions: supporting and suspending organelles, facilitating growth and expansion, protection from damage.

• Rough endoplasmic reticulum – found in eukaryotes. Functions: synthesis of proteins, protein folding and modification, transport of proteins, manufacturing or membranes

• Smooth endoplasmic reticulum – found in eukaryotes. Functions: synthesize and store lipids, produce and distribute cellular products, metabolizing carbs, detoxifying drugs, storing calcium in muscle cells

• Vacuole – in plant and fungal cells. Functions: water storage, waste disposal, maintain cell turgor, regulate pH, protect cell, buoyancy

• Golgi body – in eukaryotic cells. functions: processes and packages proteins and lipids

• Centriole – in animal and lower plant cells. Func: organizes microtubes that are used as the cell’s skeletal system

• Lysosome – in animal cells. Func: degrades molecules of nutrient or foreign particles taken from outside the cell.

• Cytoskeleton – in nearly all cells. Func: helps the cell maintain its shape and internal organization.

• Peroxisome – in eukaryotes. Func: carry out oxidative reactions in a cell using molecular oxygen.

• Central vacuole – in plant cells. Func: regulates cytoplasm compositions, stores cell compounds, maintain turgor pressure

• Flagella: in prokaryotes and eukaryotes. A flagellum is a microscopic hair-like organelle used by cells and microorganisms for movement. Flagella are filamentous protein structures found in bacteria, archaea, and eukaryotes, though they are most commonly found in bacteria. Some eukaryotic cells use flagellum to increase reproduction rates. Other eukaryotic and bacterial flagella are used to sense changes in the environment, such as temperature or pH disturbances.

Differences Between Organelles

• Cell Wall vs. Cell Membrane: The cell wall is rigid and found only in plants, while the cell membrane is flexible and present in all cells.

• Eukaryotic vs. Prokaryotic Organelles: Eukaryotic cells contain membrane-bound organelles, while prokaryotic cells do not.

• Unique Organelles: Plant cells have chloroplasts and a large central vacuole, while animal cells have centrioles and lysosomes.

Cell Membranes

Structure of Cell Membranes

• Fluid Mosaic Model: Describes the cell membrane as a flexible structure with various proteins embedded in or attached to the phospholipid bilayer.

• Phospholipid Bilayer: Composed of hydrophilic phosphate heads and hydrophobic fatty acid tails, creating a barrier to most water-soluble substances.

• Hydrophilic: attracted to water

• Hydrophobic: doesn’t like water

• Integral/Channel Proteins: Span the membrane, make the bilayer selectively permeable since they regulate items that enter and leave.

• Peripheral Proteins: Located on the inner or outer part of the bilayer, involved in signaling and maintaining the cell's shape.

Functions of Membrane Components

• Transport Proteins: Regulate the movement of substances across the membrane, maintaining homeostasis.

• Glycoproteins/Marker proteins: help the cells identify each other. Extra fact: glycoproteins on red blood cells decide a person’s blood type.

• Glycolipid: provides energy to cells, maintains stability, facilitates cellular recognition

• Cholesterol: Stabilizes membrane fluidity/flexibility, ensuring proper function across temperature variations.

Osmosis and Diffusion

- Solute: a substance dissolved in a solvent. Ex: salt, sugar

- Solvent: a substance that does the dissolving. (usually water)

- Solution: the homogeneous mixture of a solute dissolved in a solvent. Ex: salt water

- Diffusion: movement of solutes from areas of high concentration to areas of low concentration.

- Concentration gradient: exists when a solute is not at equilibrium; ex: area of high concentration vs. area of low concentration

- Passive transport: movement of particles into or out of the cell WITHOUT requiring energy. Ex: diffusion and osmosis

- Equilibrium: Equilibrium in cells refers to a state in which there is a balance between the movement of molecules into and out of the cell.

- Osmosis: movement of water from an area of high water concentration to an area of low water concentration through a semipermeable membrane

- Hypertonic solution: The solution outside that has a higher concentration of solute than the solution inside the cell. In this solution, water flows out by osmosis, into the solution outside the cell. Result: cell shrinks (plasmolysis)

- Hypotonic solution: the solute outside the cell has a lower concentration of solute than the solution inside the cell. Water moves into the cell by osmosis. Result: cell swells and bursts open (cytolysis). Plants prefer hypotonic solutions.

- Isotonic solution: the solutions inside and outside of the cell have the same concentration of solute. In this solution, water flows out and in at the same amount and rate. It doesn’t stop. Result: equilibrium, cell remains the same size

Passive transport across a membrane

- Simple diffusion: high to low concentration gradient. Simple diffusion is the process by which solutes are moved along a concentration gradient in a solution or across a semipermeable membrane.

- Facilitated diffusion: high to low concentration gradient. Facilitated diffusion is a form of facilitated transport involving the passive movement of molecules along their concentration gradient, guided by the presence of an integral/channel protein forming a channel. Channel proteins in facilitated diffusion: carrier, channel, and gated proteins

- Channel proteins in simple diffusion: none; uses hydrogen bonds.

- Channel proteins in facilitated diffusion: carrier, channel, and gated proteins

Active transport across a membrane

- Active transport: Active transport is the process of transferring substances into, out of, and between cells, using energy.

- Energy from ATP: used for transport. Active transport is most commonly accomplished by a transport protein that undergoes a change in shape when it binds with the cell’s “fuel,” a molecule called adenosine triphosphate (ATP).

- Concentration gradient: low to high

- Sodium potassium pump proteins: It is this gradient that allows our nerve cells to fire, creating muscle contractions, sensations, and even thoughts. Even our heart muscle relies upon these ion gradients to contract.

- Exocytosis (export from cell in vesicle): Exocytosis describes the process of vesicles fusing with the plasma membrane and releasing their contents to the outside of the cell. Exocytosis occurs when a cell produces substances for export, such as a protein, or when the cell is getting rid of a waste product or a toxin. Newly made membrane proteins and membrane lipids are moved on top of the plasma membrane by exocytosis.

- Endocytosis: In endocytosis, the cell uses proteins in its membrane to fold the membrane into the shape of a pocket. This pocket forms around the contents to be taken into the cell. The pocket grows until it is pinched off, re-forming the cell membrane around it and trapping the pocket and its contents inside the cell. These membrane pockets, which carry materials inside of or between cells, are called “vesicles.”