Lab Practical Flashcards
How many micro liters are in one milliliter?
1000
What is the difference between a stage micrometer and an ocular micrometer?
A stage micrometer is a miniature ruler on the stage of the microscope that has a known distance between the little lines. An ocular micrometer is a miniature ruler on the eye piece of a microscope, and its little lines never change in size, but they represent different lengths depending on how much magnification is occurring.
What are ocular units, and how can you convert between them and other units?
Ocular units are the units of the ocular micrometer which can be converted into “real” units based on relative sizes. A known distance on a stage micrometer can be compared to a unit of the ocular micrometer so measurements with the ocular micrometer can be put in perspective.
What is the difference between resolution and magnification?
Magnification refers to how many times larger something is being observed. It has nothing to do with how clear the image is.
Resolution refers to how clear something is, and specifically, the smallest distance between two points on a specimen that can still be distinguished as two separate entities.
How is resolution quantified?
Resolutions are identified by R values and the smaller the R value, the higher the resolution because this means you can distinguish between smaller points.
What is NA, and how does it relate to R?
NA, or numerical aperture, is a number that has to do with the range of angles that the lens can accept or emit light. This directly relates to resolution because R is directly correlated to wavelength of light / 2 NA
What is working distance, and how does it relate to magnification?
Working distance refers to the size of the area of something you can see under a microscope. When magnification is greater, working distance is smaller because you can see less of the object
What is the objective lens of a microscope?
Objective lens in a microscope is the lens that you choose to look at the specimen under. In the light microscopes we use, the different objective lenses are coded by different colors and have different magnification values written on them. The longer the objective lens, the higher the magnification and the smaller the working distance
What are the three main ways of creating contrast in light microscopy?
Cell staining, changing the optics (bright field vs. phase contrast), and fluorescence microscopy
What is a chromophore?
A chromophore is a chemical compound that absorbs light
What makes something a “Bradford Assay”
Making a standard curve using CBBG (coomassie brilliant blue G-250) as a dye to bind to a protein of interest. When CBBG binds to a protein, it is in de-protinated blue form with a max absorbance of 595nm
What is the absorbance equation?
Absorbance = Extinction coefficient * path length of light * concentration
A = E b c
In the Bradford Assay experiment, what protein was analyzed?
The protein was BSA (bovine serum alminum)
After centrifuging blood for the first time, what are the three layers and what are they made of?
Top - 55% water, proteins, nutrients
Middle - 1% buffy coat (white blood cells)
Bottom - 44% red blood cells
What are the two methods for precipitating proteins from a solution, and how do they work?
Salting out - salting out is adding a large amount of salt to a solution with proteins which causes the water molecules to come off of the protein so the protein can aggregate together and come out as a solid at the bottom
Organic solvent - adding the organic solvent is adding a non-polar substance to the solution with the protein with it the protein-protein interactions are stringer than the protein-solvent interactions
What does SDS stand for?
Sodium dodecyl sulfate
What does SDS do?
SDS linearizes proteins by adding a negative charge all around ten so there is an even charge to mass ratio
What is the equation for electrophoresis, and what do the variables stand for?
Ez = fv
E – Electric fileld
z – Net charge on molecule
f – frictional coefficent
v – velocity
What is bromophenol blue for?
Bromophenol blue tracts sample migration / the progress of electrophoresis but does not bind to proteins
What is coomassie blue for?
Coomassie blue binds proteins and can stain individual proteins in a gel after electrophoresis
What is the difference between agarose gel electrophoresis, and polyacrylamide gel electrophoresis?
Agarose gel electrophoresis is used for separating proteins
Polyacrylamide gel electrophoresis is used for separating DNA and RNA
