6.1-Biologists use microscopes + biochemistry to study cells

• Cell


  An organism's basic unit of structure and function
• Light microscope


  Focuses visible light passed through specimen then through glass lenses

  lenses refract (bend) the light so that the image of the specimen is magnified as it is projected into the eye or a camera
• what are the 3 important parameters in microscopy?

  magnification, resolution, and contrast
• what is magnification? how is this seen in light microscopy?

  ratio of an object’s image size to its real size

  LM can effectively magnify about 1,000x the actual size of the specimen
• at greater magnifications, can additional details be seen?

  no: cannot be seen clearly
• what is resolution?

  a measure of the clarity of the image; the minimum distance between two points can be separated and still be distinguished as separate points.

  For example, what appears to the unaided eye as one star in the sky may be resolved as twin stars with a telescope, which has a higher resolving ability than the eye.
• what is contrast? what are the methods for enhancing contrast?

  difference in brightness between the light and dark areas of an image.

  Methods for enhancing contrast include staining or labeling cell components to stand out visually
• what prevented cell biologists from using standard LM to study organelles?

  resolution barrier
• what are organelles?

  membrane-enclosed structures within eukaryotic cells
• what is the electron microscope and how it is different to light microscopes?

  focuses a beam of electrons through the specimen or onto its surface
• resolution is inversely related to light wavelengths (electrons) a for microscopic imaging
• electron beams have much shorter wavelengths than visible light.
• what is SEM? how is it undergone?

  scanning electron microscope - useful for detailed study of the topography of a specimen; an electron beam scans surface of the sample, usually coated with a thin film of gold. The beam excites electrons on surface, and these secondary electrons are detected by a device that translates the pattern of electrons into an electronic signal sent to a video screen. The result is an image of the specimen’s surface that appears three-dimensional
• TEM

  Transmission electron microscope
• What TEM is used to study

  The internal structure of cells
• How TEM works

  1. Aims an electron beam through a very thin section of the specimen

  2. Specimen has been stained with atoms of heavy metals, which attach to certain cellular structures, thus enhancing the electron density of some parts of the cell more than others

  3. Electrons passing through the specimen are scattered more in the denser regions, so fewer are transmitted

  4. The image displays the pattern of transmitted electrons
• TEM aims to study the internal structure of cells in the same way a light microscope aims light through a sample on a slide
• instead of what LM uses, what do SEM and TEM use to focus the images of cells?

  instead of glass lenses, they use electromagnets as lenses to bend the paths of the electrons, ultimately focusing the image onto a monitor for viewing.
• what did electron microscopes help reveal?

  SEM and TEM have revealed many subcellular structures that were impossible to resolve with the light microscope.
• what did the light microscope help study?

  living cells
• what is the disadvantage of electron microscopy?

  methods customarily used to prepare the specimen kills the cells and can introduce artifacts (structural features seen in micrographs that do not exist in the living cell).
• what is super-resolution microscopy?

  new imaging techniques and can label molecules instead of using markers
• what is a great advantage of super-resolution microscopy?

  allowed researchers to “break” the resolution barrier and distinguish subcellular structures as small as 10–20 nm across.
• what is the recently developed TEM?

  cyro-electron microscopy (cryo-EM)
• what does cryo-EM allow?

  allows specimens to be preserved at extremely low temperatures, avoiding the use of preservatives, and allowing visualization of structures in their cellular environment.
• where is it most used?

  method increasingly used to complement X-ray crystallography in revealing protein complexes and subcellular structures like ribosomes.

  also used to reveal some individual proteins
• cytology

  study of cell structure
• metabolism

  chemical processes of cells
• what is a useful tool for cytology?

  microscopes
• what is a useful tool for studying cell structure and function?

  cell fractionation
• what is done in cell fractionation?

  takes cells apart and separates major organelles & other subcellular structures from one another, using the equipment centrifuge
• how does a centrifuge work? what is the process called?

  spins test tubes holding mixtures of disrupted cells at a series of increasing speeds. this is called differential centrifugation
• at each speed during centrifugal, the resulting force causes a subset of cell components to settle to the bottom of the tube, forming a pellet
• at lower speeds during centrifugal, the pellet consists of larger components