lab 1 Flashcards
what is a Biuret Reagant?
alkaline solution solution complaining copper. The light absorbing center of the Biuret reagent forms a complex between two or more peptide bonds and copper ions.
what colour does the buret Reagant turn when it comes in contact with protein?
it goes from blue to purple
what is the maximal amount of light absorbed by the chromophore?
540nm
what are the 2 different ways to determine the presence of certain amino acids?
using their relative abundance as a measure of protein content and dye binding
what does a spectrophotometer do?
emits wavelengths that are then absorbed by the substance (at a maximum), which then shows the substances concentration. The spectrometer produces the light of the wavelength and the photometer measures the intensity of light by measuring the amount of light that passes through the sample.
what does BSA stand for?
bovine serum albumin
what is the concentration of BSA?
5mg/ml
how does the Biuret test allow us to detect protein?
The presence of protein in the assay will be detectable as the blue Biuret solution becomes a purple colour.
how does the Biuret test allow us to detect protein concentration in ALL material?
The intensity of the purple colour can be measured and used to estimate the concentration of proteins in a given sample, since peptide bonds occur with approximately the same frequency per gram of material in most proteins.
what is BSA? What’s it’s purpose in this lab?
the solution containing a known protein concentration, which will help us create a standard protein curve
what exactly are we doing in this lab?
1) we are making a protein standard curve for bovine serum albumin, in order to see what protein concentrations are linked to specific wavelengths. Since this applies to all proteins (a specific wavelength, for all proteins, corresponds to a specific protein concentration), we are using this information in order to determine the concentration of proteins in an unknown sample
what does our standard protein curve graph look like?
during the exam, a graph will be given to you. how will you use this graph to find protein concentration?
In class your graphs were made by hand and not with excel. For the lab exam you will also be given
a hand drawn graph that does not have an equation. You can choose to make the necessary calculations to generate an equation and determine the unknown concentrations using the equation or you can use the graph directly (using drop down lines) to determine the unknown concentration. Either would be acceptable.
what is the chromophore?
light absorbing complex of biuret Reagant
what does a darker purple colour signify?
more protein present
