Chapter 3: A tour of the cell Flashcards
What is the difference between Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM)?
- SEM uses the application of a metallic layer onto a sample which deflects the electron beam onto the detector and is used to study the external cell surface.
-TEM passes the electron bean through a thin sample and can be used to study internal cell structure.
What are the most common reagents used for protein and lipid fixing in TEM?
Protein fixing: Glutaraldehyde
Lipid fixing: Osmium tetroxide
Why are heavy metals used for sample staining in TEM?
Heavy metals preferentially bind to specific molecules and increase the contrast of images by preventing electrons passing through due to their large size (causing darker areas)
What are the two most common reagents used for sample staining in TEM and what do these elements bind to?
- Uranium: Nucleic acids and proteins
-Lead: Lipids
What piece of equipment is used to cut ultrathin EM sections?
Ultramicrotome
What is the approximate thickness of a sample suitable for TEM?
How does this compare to the thickness on EM samples?
- approx. 70 nm thick
-Much smaller than the 5-50 µm samples used in EM.
What does the electron beam hit and what occurs to make the visible image in TEM?
The electron beam hits a fluorescent screen that emits visible light on impact.
Why is the interior of an electron microscope a vaccuum?
To prevent scattering of the electron beam on impact with air molecules.
What controls the direction of the electron beam in an electron microscope?
Magnets
What is Immunoelectron microscopy?
The use of immunolabelling to localise specific molecules to particular organelles using an electron microscope.
What is cell fractionation?
A technique using disruption and ultracentrifugation of a cell/s to separate its components
When cells are disrupted (i.e. before centrifugation) what is formed?
A cell homogenate
In centrifugation, what does the rate of settlement depend on?
- Particle size, shape and density
- Liquid density
- Rotational speed of centrifuge
What is the name for the particles that have collected together at the bottom of a tube during centrifugation?
A pellet
Describe the process of differential sedimentation.
Centrifugation taking place at increasingly higher speeds to separate particles according to their size and density. Each pellet is removed from the supernatant (remaining suspension) before the next speed increase.
Describe density gradient centrifugation
A cell lysate is laid upon a pre-prepared density gradient sucrose solution. Movement through the density gradient optimises separation into bands and can be removed as fractions with a pipette or pierced hole.
How does the nucleoid of a prokaryote appear under an electron microscope - light or dark?
Light
What shape does the DNA molecule form in a prokaryote?
Circular
What is the function of glycogen granules within prokaryotic cells?
Energy storage
What does a gram-negative bacteria have that a gram-positive bacteria does not?
Periplasmic space separating the inner and outer membrane
In bacteria, what is the cell wall composed of?
Peptidoglycan (polymer of sugars linked by amino acids)
What is the difference between gram-positive and gram-negative bacteria?
Gram positive
- thick layer of peptidoglycan at surface
Gran-negative
- Thinner layer of peptidoglycan within the periplasmic space (separates inner and outer membrane)
How can Gram stain be used to distinguish and how?
Gram stain can be used to distinguish between gram-positive and gram-negative bacteria.
Gram stain is absorbed and held by gram-positive bacteria’s thick peptidoglycan layer
Gram stain is not absorbed and held by gram-negative bacteria’s thin peptidoglycan layer
After staining the gram stain is dissolved to dissolve cell membranes and decolourise the cell. Gram positive bacteria will present as purple.
What do the capsules that some bacteria possess protect them from?
Desiccation and phagocytosis by other cells
What is the purpose of a bacterium’s cell wall?
- Support
-Shape
-Prevents expansion from water absorption
Peripheral proteins and Integral membrane proteins interact with what part of the phospholipid bilayer?
Peripheral proteins - hydrophilic heads
Integral membrane proteins - hydrophobic tails
Out of Plant, Animal, Protist, and Fungal cells - which have cell walls?
Plant cells DO
Fungal cells DO
Animal cells DO NOT
Protist cells SOMETIMES DO
What does the cell membrane of eukaryotic cells contain?
Phospholipid bilayer
Proteins and glycoproteins
Cholesterol
What types of proteins and glycoproteins are related to the cell surface and what are their functions?
Transporters and channels - selective movement of ions, nutrients, and other molecules across the membrane
Receptors - Responding to molecular changes in the extracellular environment
Recognition molecules - Promote adhesion between adjacent tissue cells
Intracellular proteins - Attached to cytoplasmic surface (includes cytoskeleton with structural roles)
What are the three protein assemblies that form the cytoskeleton of a eukaryotic cell and what are their constituents?
Microfilaments (actin filaments) - Actin
Intermediate filaments - Several different constituents
Microtubules - α and β Tubulin
What are the approximate diameters of Microfilaments, Microtubules, and intermediate filaments?
Microfilaments - 6 nm
Microtubules - 25 nm
Intermediate filaments - 8 - 10 nm
What is the structure of microfilaments?
Long helical chains of actin linked together in networks or bundles
What is the structure of microtubules?
Hollow tubes made up of (usually) 13 parallel filaments of tubulin assemblies
What is the structure of Intermediate filaments?
Rope-like structures of approximately 8 protofilaments wound around eachother
Where are each of the cytoskeleton filaments most prominent within a eukaryotic cell?
- Microfilaments | In the cell cortex, within microvilli, the leading edge of moving cells
- Microtubules | Radiating from the centrosome (itself near the nucleus)
- Intermediate filaments | All over (links to adjacent cells and extracellular membrane)
What is the function of microfilaments and how does their abilities facilitate this?
Their ability to quickly disassemble and reassemble makes it successful as a tool for cell motility.
What are the functions of microtubules?
- Cell shape maintenance
- Intracellular movement
- Reorganisation of chromosomes during cell division
- Role in distribution of organelles
- Components of cilia and flagella
Are microtubules stable or unstable? What can change this?
Extremely unstable.
Stabilised when it attaches to a membrane, organelle, or capping protein.
Intermediate filaments have what functions?
- Provide mechanical strength to cells and tissues (they are strong)
- Link adjacent cells and extracellular matrix
What occurs within the nucleus within a cell?
DNA replication
Transcription of DNA into RNA
Ribosomal assembly
What regulates the nuclear structure (Cell nucleus)?
What do the components link to?
Nuclear Lamina (network of a type of intermediate filament - lamins)
They link to the nuclear membrane and the chromatin
What are the components of the eukaryotic cell nucleus?
- Nuclear envelope
-Nuclear pores
- Nuclear pore complex (complex protein arrangements)
- Nucleolus
- Genetic material
What is the approximate size of nuclear pores and what do they allow?
Approx. 9 nm
Allow free movement of water molecules
What is DNA wound round?
Histones (proteins)
What is the difference between heterochromatin and euchromatin, where they are found within the nucleus, and how they appear on EM?
- Heterochromatin is tightly packed DNA, has fewer genes, more prominent near the nuclear envelope, and appears dark on EM.
-Euchromatin is loosely packed DNA, has more genes, found more central in the nucleus, appears pale on EM.
Why is some DNA packed more densely or loosely within a cellular nucleus?
As part of gene expression. Less densely packed DNA is able to be accessed by enzymes involved in transcription.