Chapter 2

• Magnification 


  factor by which image is larger than actual specimen
• Resolution 


  smallest distance at which 2 seperate structures can be distinguished from one another
• Dry mount method 


  • solid specimens viewed as whole or cut into very thin slice w a sharp blade - sectioning
  • specimen placed on centre of slide & coverslip placed over sample
• Wet mount method 


  • specimen suspended in liquid (water/ immersion oil)
  • coverslip placed on from an angle
• Squash slides 


  • wet mount first prepared
  • lens tissue used to gently press down coverslip
  • care taken so coverslip not broken when being pressured
• Smear slides


  • edge of slide used to smear sample - create thin even coating on another slide
  • cover slip placed over sample
• Why do we need to stain cells?

  • samples often have low contrast as they don't absorb a lot of light
  • stain increases contrast as diff parts in cell take up stains to diff degrees - allows components to become visible so they can be identified
• Positive staining 


  • crystal violet or methylene blue - positive charge
  • attracted to negatively charged cell structures - lead to staining
• Negative staining 


  • nigrosin or congo red - negative charge
  • repelled by the negative charged cytosol - dyes stay outside of cells, leaving cell unstained
  • cells stand out against stained background
• Differential staining

  can distinguish between structures in cells or between different species that would otherwise be hard to identify
• Gram-positive bacteria


  susceptible to penicillin - inhibits formation of peptidoglycan cell walls
• Gram-negative 


  thinner walls, not susceptible to penicillin
• Gram staining 


  • crystal violet applied - iodine added to fix dye - slide washed w alcohol
  • gram-positive retain stain - appear blue/purple
  • gram-negative - thin walls - lose stain; stained w counterstain (safranin dye) - appear red
• Carbolfuchsin staining (acid-fast technique) 


  • lipid solvent used to carry carbolfuchsin dye into cells
  • cells washed w dilute acid-alcohol solution
  • mycobacterium not affected by acid-alcohol - retain stain - appear red
  • other bacteria lose stain - exposed to methylene stain - appear blue
• How does a light microscope work?

  • light shines up through the sample being observed & through the 2 lenses (objective & eyepiece), each lens magnifies the image
  • magnification up to x2000
• Why do electron microscopes have more detail than light microscopes?

  • electron beams have shorter wavelengths than light waves
  • so electron microscopes have greater resolution
• Scanning electron microscope (SEM)

  • beam of electrons is sent across the surface of a specimen & reflected electrons are collected
  • resolution not as high as TEM - 3-10nm
  • magnification up to x100,000
  • produces 3D images
• Transmission electron microscope (TEM)


  • beam of electrons is transmitted through a specimen
  • best resolution - 0.5nm
  • magnification up to x500,000
  • enables intracellular details to be observed
• Light microscope vs electron microscope

  • inexpensive/ expensive
  • small & portable/ large & needs to be installed
  • simple sample prep/ complex prep
  • sample preparation doesn't usually lead to distortion/ does distort material (produce artefact)
  • vacuum not required/ vacuum required
  • natural colour/ black & white
  • specimens living or dead/ specimens dead
  • magnification up to x2000/ x500,000
• What's an artefact?


  structure that is produced due to the preparation process that is not a natural part of the specimen
• Structure of the nucleus

  • contained within a double membrane called nuclear envelope - protect from damage in cytoplasm
  • nuclear envelope contains nuclear pores - allow substances to enter/ exit
  • nucleolus - composed of protein & RNA - forms ribosomes need for protein synthesis
• Nucleus function 


  • contains DNA - DNA associates w proteins called histones to form chromatin which coils & condenses to form chromosomes
  • controls: gene expression, site of mRNA transcription & mitosis
• Structure of smooth endoplasmic reticulum (SER)

  network of flattened membrane-bound sacs called cisternae that connect to nuclear envelope
• Function of SER 


  lipid & carbohydrate synthesis, storage
• Structure of rough endoplasmic reticulum (RER)

  network of cisternae bound to ribosomes
• Function of RER 


  protein synthesis & transport
• Structure of ribosomes 


  • can be free-floating or attached ER to form RER
  • formed of RNA & proteins
  • have large subunit - joins amino acids
  • small subunit - mRNA binding site
• Function of ribosomes 


  site of protein synthesis
• Structure of Golgi apparatus 


  compact structure formed of cisternae
• Function of Golgi apparatus

  modifies & packages proteins into vesicles
• Structure of mitochondria 


  • double membrane
  • inner membrane - folds to form cristae
  • internal fluid called matrix

  
• Function of mitochondria 


  • ATP production through aerobic respiration
  • more mitochondria in cell = more active cell
• Structure of vesicles 


  • membranous sacs
  • single membrane with fluid inside
• Function of vesicles 


  transport materials inside cell
• Structure of lysosomes 


  specialised vesicles containing hydrolytic enzymes
• Function of lysosomes 


  work in immune system - break down waste material/ pathogens
• Structure of centrioles 


  • composed of microtubules
  • 2 associated centrioles form the centrosome
• Function of centrioles 


  involved in assembly & organisation of spindle fibres during cell division
• Importance of the cytoskeleton

  • microfilaments - control cell movement & cell contraction during cytokinesis
  • microtubules - regulate shape & organelle movements. form centrioles & spindle fibres
  • intermediate fibres - give mechanical strength to cells
• Protein production 


  • mRNA copy made in nucleus - leaves through nuclear pore
  • mRNA attaches to a ribosome - bound to RER - ribosomes synthesise protein
  • protein 'pinched off' in vesicles & travel to golgi
  • golgi modifies & packages protein - protein leaves on vesicle
  • secretory vesicles carry proteins that are to be released - moves towards & fuse w plasma membrane & contents released
  • some vesicles form lysosomes - contain enzymes for use in cell