Welcome to Cell Ultrastructure and Microscopy!
Have you ever wondered what goes on inside the microscopic building blocks of life? Just like a bustling city has power stations, roads, waste recycling centres, and a town hall, living cells have specialised internal structures called organelles that keep everything running smoothly.
In this chapter, we will explore the internal architecture of cells (their ultrastructure), compare complex eukaryotic cells with simpler prokaryotic cells, trace how proteins are produced and exported, and master the microscopy tools and mathematical calculations you need for your exams.
Note: Detailed plant-specific structures (such as chloroplasts and amyloplasts) are explored in our dedicated chapter on Plant Cells, Fibres and Sustainability.
1. Levels of Structural Organisation
Before diving inside a single cell, let's look at how living things are assembled from the ground up. In multicellular organisms, cells do not work in isolation; they coordinate across different levels of hierarchy:
Cells \(\rightarrow\) Tissues \(\rightarrow\) Organs \(\rightarrow\) Organ Systems \(\rightarrow\) Organism
Let's break down each level with clear definitions:
1. Cell: The fundamental structural and functional unit of life (for example, an epithelial cell or a muscle cell).
2. Tissue: A group of similar, specialised cells working together to perform a specific function (for example, epithelial tissue lining organs, or muscle tissue contracting to cause movement).
3. Organ: A group of different tissues working together to carry out a particular overall function (for example, the stomach, which contains epithelial tissue, muscle tissue, and glandular tissue).
4. Organ System: A group of organs working cooperatively to carry out major life processes (for example, the digestive system or the cardiovascular system).
Key Takeaway: Cells group into tissues, tissues group into organs, and organs group into organ systems to keep an organism alive and functioning.
2. Eukaryotic Cell Ultrastructure
The word ultrastructure refers to the fine, detailed internal architecture of a cell that can only be revealed using high-resolution electron microscopes.
Eukaryotic cells are cells that contain a true, membrane-bound nucleus and membrane-bound organelles. Let's tour the key eukaryotic organelles required by your syllabus:
The Nucleus and Nucleolus
Structure: Surrounded by a double membrane called the nuclear envelope, which is pierced by small channels called nuclear pores. Inside, it contains chromatin (DNA wrapped around histone proteins) and a dense region called the nucleolus.
Function: The nucleus controls all cellular activities by housing the genetic instructions (DNA). The nuclear pores allow the transport of molecules (such as mRNA leaving the nucleus). The nucleolus is the specific site where ribosomes are assembled.
Ribosomes
Structure: Tiny, non-membrane-bound particles made of ribosomal RNA and protein. In eukaryotic cells, these are 80S ribosomes (which are larger than the 70S ribosomes found in prokaryotes). They can float freely in the cytoplasm or be bound to the outer surface of the rough endoplasmic reticulum.
Function: The site of translation during protein synthesis.
Rough Endoplasmic Reticulum (rER)
Structure: An extensive network of flattened, membrane-bound sacs (called cisternae) studded with ribosomes on its outer surface. It is often continuous with the outer nuclear membrane.
Function: Folds and packages newly synthesised proteins into transport vesicles to be shuttled to the Golgi apparatus.
Smooth Endoplasmic Reticulum (sER)
Structure: Similar to the rER with interconnected tubular membrane sacs, but it completely lacks ribosomes on its surface.
Function: Synthesises, stores, and transports lipids and steroids (such as cholesterol).
Golgi Apparatus
Structure: A stack of flattened, curved, membrane-bound sacs called cisternae, surrounded by small spherical vesicles. Unlike the ER, the cisternae of the Golgi are not physically interconnected.
Function: Modifies proteins received from the rER (for example, by adding carbohydrate chains to form glycoproteins) and packages them into vesicles for transport to other parts of the cell or for secretion outside the cell (exocytosis). It also forms lysosomes.
Lysosomes
Structure: Spherical, single-membrane-bound vesicles containing powerful digestive hydrolytic enzymes (lysozymes).
Function: Break down worn-out organelles, debris, and foreign material engulfed by the cell (e.g. during phagocytosis by white blood cells).
Mitochondria (Singular: Mitochondrion)
Structure: Enclosed by a double membrane. The outer membrane is smooth, while the inner membrane is folded into finger-like projections called cristae. The cristae enclose an inner fluid-filled space known as the matrix, which contains circular mitochondrial DNA and 70S ribosomes.
Function: The site of aerobic respiration, producing ATP to supply energy for cellular activities.
Centrioles
Structure: Small, hollow cylinders made of a ring of nine triplets of microtubules. In animal cells, they occur in pairs located just outside the nucleus in a region called the centrosome.
Function: Involved in the assembly and organisation of the spindle fibres during nuclear division (mitosis and meiosis).
Memory Aid for Double Membranes: Remember "NM" (Nuclear envelope, Mitochondria) are the organelles with double membranes in animal cells!
Key Takeaway: Eukaryotes partition cellular tasks into membrane-bound compartments (organelles), allowing incompatible chemical reactions to happen simultaneously without interference.
3. The Pathway of Protein Transport and Extracellular Enzymes
One of the most frequently examined topics is how cells manufacture and export proteins, such as extracellular enzymes (enzymes that work outside the cell, like digestive enzymes). Don't worry if this seems like a lot of steps—it works just like an automated factory assembly line!
Step-by-Step Pathway:
1. Transcription in the Nucleus: The gene coding for the protein is transcribed into messenger RNA (mRNA). The mRNA exits the nucleus through a nuclear pore.
2. Translation on the rER: Ribosomes attached to the rough endoplasmic reticulum translate the mRNA into a polypeptide chain. The growing polypeptide moves into the lumen of the rER, where it folds into its 3D secondary and tertiary structure.
3. Transport Vesicle Formation: The folded protein is packaged into a small transport vesicle that pinches off (buds) from the rER.
4. Transport to the Golgi: The vesicle moves through the cytoplasm and fuses with the receiving face of the Golgi apparatus.
5. Modification and Sorting: As the protein moves through the Golgi cisternae, it is modified (e.g. carbohydrates added) and sorted for export.
6. Secretory Vesicle Formation: The finished protein is packaged into a secretory vesicle that pinches off from the Golgi apparatus.
7. Exocytosis: The secretory vesicle travels to the cell surface membrane, fuses with it, and releases the extracellular enzyme outside the cell.
Exam Tip: When exam questions ask you to describe protein transport, be sure to clearly mention the roles of both transport vesicles (from rER to Golgi) and secretory vesicles (from Golgi to cell surface membrane)!
4. Prokaryotic Cell Ultrastructure
Prokaryotes (such as bacteria) are unicellular organisms that are much smaller and simpler than eukaryotes. The word prokaryote literally means "before a nucleus".
Prokaryotic Features and Their Roles:
1. Cell Wall: A rigid external layer made of peptidoglycan (murein) that prevents the cell from bursting due to osmotic pressure and maintains cell shape.
2. Capsule: A protective slimy layer outside the cell wall that prevents the bacterium from drying out and helps shield it from attacks by the host's immune system.
3. Circular DNA: A single, long, circular loop of double-stranded DNA that floats freely in the cytoplasm in a region called the nucleoid. It is naked (not bound to histone proteins).
4. Plasmids: Small, separate circular loops of DNA that can replicate independently. They often carry non-essential but advantageous genes, such as antibiotic resistance genes, and can be transferred between bacteria.
5. Ribosomes: 70S ribosomes, which are smaller than the 80S ribosomes found in the cytoplasm of eukaryotic cells, used for protein synthesis.
6. Flagellum (Plural: Flagella): A rigid, rotating helical protein filament that acts like a propeller, allowing the bacterium to move.
7. Pili (Singular: Pilus): Thin, hair-like protein projections on the cell surface that help bacteria attach to host tissues or to other bacteria during the transfer of plasmids.
Comparing Eukaryotic and Prokaryotic Cells
Exam questions frequently ask you to compare and contrast prokaryotes and eukaryotes. Remember: in Edexcel exams, "compare and contrast" requires you to state both similarities and differences!
Similarities:
- Both possess a cell surface (plasma) membrane.
- Both contain ribosomes for protein synthesis.
- Both contain genetic material made of DNA.
Differences:
- Nucleus: Prokaryotes have no nucleus (DNA is free in the cytoplasm as circular DNA); eukaryotes have a membrane-bound nucleus containing linear DNA.
- Membrane-bound organelles: Prokaryotes completely lack membrane-bound organelles (no mitochondria, ER, or Golgi); eukaryotes have membrane-bound organelles.
- Ribosomes: Prokaryotes have smaller 70S ribosomes; eukaryotic cytoplasm has larger 80S ribosomes.
- Cell wall: When present in prokaryotes, the wall is made of peptidoglycan; animal cells have no cell wall at all.
- DNA organisation: Prokaryotic DNA is circular and naked; eukaryotic nuclear DNA is linear and associated with histone proteins.
Key Takeaway: Prokaryotes are simpler, lack a nucleus and membrane-bound organelles, have circular naked DNA and 70S ribosomes, and may possess specialised structures like plasmids, capsules, pili, and flagella.
5. Microscopy: Magnification, Resolution, and Staining
To view microscopic cells and their tiny organelles, biologists rely on two primary types of microscopes: light microscopes and electron microscopes.
Magnification vs. Resolution
These two terms are often confused by students, but they mean completely different things:
Magnification: How many times larger the image is compared to the actual size of the object.
Resolution: The minimum distance between two distinct points that allows them to still be seen as two separate entities. In simple terms, resolution is all about image detail and clarity.
Analogy: Imagine zooming in on a low-resolution digital photo. You can magnify it as much as you like, but past a certain point, the image just becomes blurry pixels. Magnification makes things bigger; resolution lets you see fine detail!
Comparing Light and Electron Microscopes
Light Microscope:
- Uses a beam of light focused by glass lenses.
- Relatively low magnification and lower resolution (limited by the longer wavelength of light).
- Organelles visible: Nucleus, cytoplasm, cell membrane, cell wall, and mitochondria (only as tiny dots).
- Can view living or dead specimens in natural colour.
Electron Microscope:
- Uses a beam of electrons focused by electromagnets.
- Very high magnification and much higher resolution (because the wavelength of electrons is thousands of times shorter than light).
- Organelles visible: Fine ultrastructure including ribosomes, internal cristae of mitochondria, endoplasmic reticulum, and Golgi apparatus.
- Specimens must be viewed in a vacuum, meaning only dead/fixed material can be examined. Images are originally black-and-white (monochrome).
The Role of Staining
Most biological materials are transparent and colourless. Staining is used to increase contrast between different cell structures, making them visible under the microscope:
- In light microscopy, coloured stains bind specifically to certain chemicals (e.g., methylene blue stains DNA blue).
- In electron microscopy, heavy metal salts (like lead or uranium) are used to scatter electrons and create contrast in micrographs.
Recognising Organelles in Electron Micrographs
In written papers, you will often be given an electron micrograph and asked to identify organelles. Here is how to recognise them quickly:
- Nucleus: Large, round, grey organelle, often with a prominent dark circle inside (the nucleolus).
- Mitochondrion: Sausage- or oval-shaped with clear internal folded membranes (cristae).
- Rough Endoplasmic Reticulum (rER): Parallel lines or sheets of membranes covered with tiny black dots (ribosomes).
- Golgi apparatus: Curved, smooth membrane stacks with circular vesicles budding off at the edges (no black dots on the membranes).
- Lysosome: Dark, uniform, single-membrane-bound circular vesicle containing enzymes.
Key Takeaway: Electron microscopes provide much higher resolution than light microscopes because electrons have a much shorter wavelength than light. Stains provide the contrast needed to see transparent cell structures.
6. Calculations: Magnification and Scale
Under the Edexcel syllabus, calculating magnification and actual sizes is a fundamental mathematical skill tested in both Unit 2 and Unit 3 papers.
The Magnification Formula Triangle
The relationship between magnification, image size, and actual size is given by the formula:
\( \text{Magnification} = \frac{\text{size of image}}{\text{size of real object}} \)
We often write this as:
\( M = \frac{I}{A} \)
You can rearrange this formula depending on what you need to calculate:
- To find actual size: \( A = \frac{I}{M} \)
- To find image size: \( I = M \times A \)
Crucial Step: Converting Units
The single most common mistake students make is using different units for image size and actual size! You must always convert both measurements to the same unit before doing the calculation.
The units you need to master are millimetres (\( \text{mm} \)) and micrometres (\( \mu\text{m} \)):
\( 1\text{ mm} = 1000\text{ }\mu\text{m} \)
- To convert from \( \text{mm} \) to \( \mu\text{m} \): multiply by \( 1000 \).
- To convert from \( \mu\text{m} \) to \( \text{mm} \): divide by \( 1000 \).
Worked Example: Calculating Actual Size
Question: An electron micrograph shows a mitochondrion with an image length of \( 48\text{ mm} \). The magnification is stated as \( \times 16000 \). Calculate the actual length of the mitochondrion in micrometres (\( \mu\text{m} \)).
Step 1: Write down what you know and convert units to \( \mu\text{m} \).
Image size (\( I \)) = \( 48\text{ mm} = 48 \times 1000 = 48000\text{ }\mu\text{m} \)
Magnification (\( M \)) = \( 16000 \)
Step 2: State the formula.
\( A = \frac{I}{M} \)
Step 3: Substitute the numbers and calculate.
\( A = \frac{48000\text{ }\mu\text{m}}{16000} = 3\text{ }\mu\text{m} \)
Final Answer: The actual length of the mitochondrion is \( 3\text{ }\mu\text{m} \).
Using a Scale Bar
Sometimes an exam question gives you a scale bar instead of stating the magnification directly. Don't panic! Here is the foolproof 3-step method:
1. Measure the length of the scale bar using a ruler in millimetres (this is your \( I_{\text{bar}} \)).
2. Read the actual size stated on the scale bar and convert both to the same units (this is your \( A_{\text{bar}} \)).
3. Calculate magnification: \( M = \frac{I_{\text{bar}}}{A_{\text{bar}}} \). Once you have \( M \), you can find the actual size of any structure in the image using \( A = \frac{I}{M} \).
7. Core Practical 5: Light Microscope Observations
Core Practical 5: Light microscope observations and labelled drawings of animal cells, with a graticule for measurement and scale.
The Apparatus: Eyepiece Graticule and Stage Micrometer
To measure cells accurately under a light microscope, you need two measuring tools:
1. Eyepiece Graticule: A transparent glass disc with an arbitrary ruler scale (typically marked from \( 0 \) to \( 100 \) divisions) placed inside the microscope eyepiece. The divisions do not have absolute units because their apparent size changes whenever you change the objective lens magnification.
2. Stage Micrometer: A microscope slide with an etched, precisely measured scale of known distance (for example, \( 1\text{ mm} \) divided into \( 100 \) small divisions, meaning each small division is exactly \( \frac{1\text{ mm}}{100} = 0.01\text{ mm} = 10\text{ }\mu\text{m} \)).
How to Calibrate the Eyepiece Graticule (Step-by-Step):
Step 1: Place the stage micrometer on the microscope stage and focus on the etched scale using the desired objective lens.
Step 2: Align the scale of the eyepiece graticule with the scale of the stage micrometer so that the zero lines on both scales coincide.
Step 3: Look along the scales to find a second point where two division lines overlap cleanly.
Step 4: Count the number of eyepiece graticule units (EGU) and the corresponding distance on the stage micrometer.
Step 5: Calculate the actual distance represented by 1 eyepiece graticule unit:
\( \text{Distance of 1 EGU} = \frac{\text{Distance on stage micrometer}}{\text{Number of eyepiece graticule units}} \)
Example: If \( 40 \) graticule units line up with \( 20 \) small divisions of a stage micrometer (where each division is \( 10\text{ }\mu\text{m} \)):
Total micrometer distance = \( 20 \times 10\text{ }\mu\text{m} = 200\text{ }\mu\text{m} \)
\( 1\text{ EGU} = \frac{200\text{ }\mu\text{m}}{40} = 5\text{ }\mu\text{m} \).
Now, if an animal cell measures \( 6 \) graticule units wide under this same lens, its actual width is \( 6 \times 5\text{ }\mu\text{m} = 30\text{ }\mu\text{m} \)!
Biological Drawings Rules
In written practical papers (Units 2 and 3), you may be asked to evaluate or produce biological drawings. Always remember these golden rules:
- Use a sharp HB pencil.
- Draw clear, unbroken, single lines (no sketching, shading, or cross-hatching).
- Ensure proportions are accurate and the drawing is large (occupying at least half the available space).
- Use a ruler to draw straight, horizontal label lines that touch the feature being identified precisely.
- Do not let label lines cross each other, and do not add arrowheads to the lines.
- Always include a scale bar and state the magnification.
Key Takeaway: Eyepiece graticules have arbitrary units and must be calibrated using a stage micrometer of known size for each objective lens before taking cell measurements.
Summary Checklist: Are You Exam Ready?
- Can you define cell, tissue, organ, and organ system?
- Can you identify eukaryotic organelles (nucleus, nucleolus, rER, sER, Golgi apparatus, mitochondria, lysosomes, centrioles, 80S ribosomes) and describe their functions?
- Can you trace a protein's journey from translation on the rER through the Golgi apparatus and into an extracellular secretory vesicle?
- Can you describe the roles of prokaryotic structures (circular DNA, 70S ribosomes, peptidoglycan cell wall, capsule, plasmids, flagellum, pili)?
- Can you state at least two similarities and three differences between prokaryotes and eukaryotes?
- Can you explain why electron microscopes have a higher resolution than light microscopes?
- Can you convert between \( \text{mm} \) and \( \mu\text{m} \) and calculate magnification using \( M = \frac{I}{A} \)?
- Can you explain how an eyepiece graticule is calibrated using a stage micrometer in Core Practical 5?