3 important parameters in microscopy:
- magnification: the ratio of an object’s image size to its real size
- resolution: measure of the clarity of the image; the minimum distance 2 points can be separated and still be distinguished as 2 points
- contrast: accentuates differences in parts of the sample
electron microscope (EM)
Rather than light, the EM focuses a beam of e- through the specimen or onto its surface. Resolution is inversely related to the wavelength of the radiation a microscope uses for imaging, and electron beams have much shorter wavelengths than visible light.
scanning electron microscope (SEM)
Useful for detailed study of the topography of a specimen. The e- beam scans the surface of the sample, usually coated w/ a thin film of gold. The beam excites e- on the surface, and these secondary e- are detected by a device that translates the pattern of e- onto an electronic signal to a video screen. The result is an image of the specimen’s surface that appears 3D.
transmission electron microscope (TEM)
Useful for studying the internal structure of cells. TEM aims an e- beam through a very thin section of the specimen. The specimen has been stained with atoms of heavy metals, which attach to certain cellular structures, thus enhancing the e- density of some parts of the cell more than others. The e- passing through the specimen are scattered more in the denser regions, so fewer are transmitted. The image displays the pattern of transmitted e-. Instead of using glass lenses, the TEM uses electromagnets as lenses to bend the paths of the e-, ultimately focusing the image on a monitor for viewing.
specimen preparation
In e- microscopy, may kill the cells.
For all microscopy techniques, specimen preparation can introduce artifacts, structural features seen in micrographs that don’t exist in the living cell.
cell fractionation
Takes cells apart and separates major organelles and other subcellular structures from one another. The centrifuge is used; it spins test tubes holding mixtures of disrupted cells at a series of increasing speeds. At each speed, the resulting force causes a fraction of the cell components to settle to the bottom of the tube, forming a pellet. At lower speeds, the pellet consists of larger components, and higher speeds yield a pellet with smaller components.
cell fractionation advantage
Enables researchers to prepare specific cell components in bulk and identify their functions, a task not usually possible w/ intact cells.
cytosol vs cytoplasm
cytosol: semifluid, jellylike portion of the cytoplasm
cytoplasm: the contents of the cell bounded by the plasma membrane; in eukaryotes, the portion exclusive of the nucleus.
all cells share certain basic features:
- bounded by plasma membrane
- cytosol
- chromosomes
- ribosomes
surface area
Cells are small b/c smaller objects have a greater ratio of SA:V.
nuclear membrane and pores
An intricate protein structure called a pore complex lines each pore and regulates the entry and exit of proteins, RNAs, and large complexes of macromolecules.
nuclear lamina
The interior side of the nuclear envelope is lined by the nuclear lamina, a netlike array of protein filaments that maintains the shape of the nucleus by mechanically supporting the nuclear envelope.
chromosome
one long DNA molecule + associated proteins, some of which help coil the DNA molecule of each chromosome, reducing its length
nucleolus
- rRNA is synthesized from instructions in DNA
- proteins imported from cytoplasm are assembled w/ rRNA into large and small subunits of ribosomes; these subunits then exit to the cytoplasm, where a large and a small subunit can assemble into a ribosome
ribosomes
- free: suspended in cytosol; make proteins that function in the cytosol
- bound: attached to the outside of ER or nuclear envelope; make proteins that are destined for insertion into membranes, packaging within certain organelles, or secretion
vesicle
A membranous sac in the cytoplasm of a eukaryotic cell. The endomembrane system is related either through physical continuity or by the transfer of membrane segments as tiny vesicles.
endomembrane system: includes
nuclear envelope, ER, Golgi apparatus, lysosomes, vesicles + vacuoles, plasma membrane
endomembrane system: function
- protein synthesis
- transport or proteins into membranes, organelles, or out of the cell
- metabolism and movement of lipids
- detoxification of poisons
endoplasmic reticulum (ER)
An extensive membranous network in eukaryotic cells, continuous with the outer nuclear membrane. Consists of a network of membranous tubules and sacs called cisternae.
functions of smooth ER
- synthesis of lipids
- metabolism of carbohydrates
- detoxification of drugs and poisons: usually involves adding hydroxyl groups to drug molecules, making them more soluble and easier to flush from the body
- storage of calcium ions
glycoprotein
A protein with one or more covalently attached carbohydrates. Most secretory proteins are glycoproteins.
functions of rough ER
- Ribosomes attached to rough ER produce secretory proteins. As a polypeptide chain grows from a bound ribosome, the chain is threaded into the ER lumen through a pore formed by a protein complex in the ER membrane. As the polypeptide enters the ER lumen, it folds into its native shape. The secretory protein departs from the ER wrapped in the membranes of vesicles that bud like bubbles from a specialized region called transitional ER.
- Membrane factory for the cell; it grows in place by adding membrane proteins and phospholipids to its own membrane.
- Produce hydrolytic enzymes (used in lysosomes) and lysosomal membrane, which are transferred to the Golgi for further processing.
Golgi apparatus
- consists of cisternae, flattened membranous sacs
- cis face: near ER; receiving department
- trans face: shipping department
- The Golgi adds molecular identification tags, such as phosphate groups. The trans face gives rise to vesicles that pinch off and travel to “docking sites,” external molecules on other organelles.
lysosome
Membranous sac of hydrolytic enzymes that an animal cell uses to digest (hydrolyze) macromolecules. Work best in the acidic environment found in lysosomes. (Ineffective in cytosol, which has neutral pH.)
