3.1 Methods Of Studying Cells Flashcards
What are microscopes?
Microscopes are instruments that produced a magnified image of an object.
Why are light microscopes have a limited resolution?
Light microscopes = long wavelengths = can only distinguish between 2 objects if they are 0.2um>= apart.
Electron microscopes = beams of electrons = shorter wavelengths = can distinguish 2 objects only 0.1nm apart.
What is magnification? Equation?
The magnification of an object is how many times bigger the image is when compared to the object.
magnification =
size of image / size of real object
What is an object? What is image?
The material that is put under a microscope is referred to as the object.
The appearance of this material when viewed under the microscope is referred to the object
Define Resolution
The resolution, or resolving power, of a microscope is the minimum distance apart that two objects can be in order for them to appear as separate items.
What is the resolution of the light microscope?
Whatever the type of microscope, the resolving power depends on the wavelength or form of radiation used.
In a light microscope it is about 0.2um.
This means that any two objects which are 0.2 μm or more apart will be seen separately, but any objects closer than 0.2 μm will appear as a single item.
In other words, greater resolution means greater clarity, that is the image produced is clearer and more precise.
What does increasing the magnification do?
Increasing the magnification increases the size of an image, but does not always increase the resolution. Every microscope has a limit of resolution.
Up to this point increasing the magnification will reveal more detail but beyond this point increasing the magnification will not do this.
The object, while appearing larger, will just be more blurred.
What is cell fractionation?
Cell fractionation is the process where cells are broken up and the different organelles they contain are separated out.
In order to study the structure and function of the various organelles that make up cells, it is necessary to obtain large numbers of isolated organelles.
What 3 things are done before cell fractionation can begin, and why?
Before cell fractionation can begin, the tissue is placed in a cold, buffered solution of the same water potential as the tissue.
The solution is:
⚫ cold- to reduce enzyme activity that might break down the organelles
⚫ is of the same water potential as the tissue- to prevent organelles bursting or shrinking as a result of osmotic gain or loss of water
⚫ buffered - so that the pH does not fluctuate. Any change in pH could alter the structure of the organelles or affect the functioning of enzymes.
What is Homogenation? (In Cell Fractionation)
Cells are broken up by a homogeniser (blender).This releases the organelles from the cell. The resultant fluid, known as homogenate, is then filtered to remove any complete cells and large pieces of debris.
What is Ultracentifugation? (in CF)
Ultracentrifugation is the process by which the fragments in the filtered homogenate are separated in a machine called a centrifuge.
This spins tubes of homogenate at very high speed in order to create a centrifugal force.
Describe the process of Cell Fractionation in detail.
The tube of filtrate is placed in the centrifuge and spun at a slow speed.
⚫ The heaviest organelles, the nuclei, are forced to the bottom of the tube, where they form a thin sediment or pellet.
⚫ The fluid at the top of the tube (supernatant) is removed, leaving just the sediment of nuclei.
⚫ The supernatant is transferred to another tube and spun in the centrifuge at a faster speed than before.
⚫ The next heaviest organelles, the mitochondria, are forced to the bottom of the tube.
⚫ The process is continued in this way so that, at each increase in speed, the next heaviest organelle is sedimented and separated out.
