Q 1. In this practical exercise you will be using biuret reagent. Why is biuret reagent used in this practical?
The biuret reagent is used to assess the concentration of the protein because peptide bonds occur with the same frequency per amino acid in the peptide. The intensity of the colour, and therefore the absorption, is directly proportional to the protein concentration.
Q 2. What is the experimental reason for constructing a standard curve?
Multiple samples with known properties can be measured and graphed, which then enables the same properties to be determined for the unknown samples, in this case the concentration, by interpolating the graph which depicts the relationship between the absorbance and concentration.
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One way in which the experimental procedure could be modified is diluting the sample. This involves decreasing the volume of BSA Protein solution and increasing the buffer volume by the same amount. As a result the absorbance would be lower and fit on the graph. Another way may be to extend the curve by increasing the concentration of the BSA Protein solution which would provide a greater range.
Q 7. Do your results support your hypothesis regarding these tissue samples? (2-3 sentences)
The hypothesis regarding the tissue samples was supported. The absorbance of the sun raised silverbeet plant was found to be 0.801, which was greater than the shaded silverbeet plant with an absorbance of 0.369.
Q 8. One possible reason for the results not supporting your experimental hypothesis is experimental error. List three examples of possible sources of error that may have occurred in your work in this practical class. (2-3 sentences)
A possible error may be adding inaccurate amounts of buffer between samples, causing them to dilute either excessively or insufficiently, resulting in an inaccurate standard curve. Another error could be leaving the sample solutions for over 25 minutes, which could cause the protein to start degrading. Not zeroing the spectrophotometer or cleaning the tube properly can affect the absorbance value.
Q 9. Even if your results do support the experimental hypothesis, give three major possible
With these absorbance numbers a concentration curve was constructed and the unknown solution was determined by finding the point of absorbance on the curve.
2. Write a statement to explain the molecular composition of the unknown solution based on the results obtained during testing with the Biuret solution and each sample solution.
The Beers Law calibration experiment used many concentrations of crystal violet solutions. Each of these solutions were test and analyzed in order to determine the absorbance of each concentration The results were than graphed and produced a slope of 1.00E05 with an intercept of -2.21E-02.
5. The degree of precision was to 3 significant figures obtained with the spectrophotometer. The major source of error in our experiment was not calibrating the spectrophotometer with distilled water.
The absorbance with SDS shows a rapid increase, before the graph begins to increase more slowly and uniformly. The absorbance levels off at 0.3nm. The absorbance with no SDS remains at a constant level of 0.1nm for the entire experiment. The absorbance of the control solution is also constant throughout the experiment. This shows the absorbance of the compounds in the solution without the ovalbumin, by taking this figure away from the other recordings, it is possible to discover the absorbance for ovalbumin alone.
To identify systematic error the experiment must be repeated. When repeating the experiment new apparatus or materials must be used, such as new stock solutions of hydrogen peroxide. If the new data differs consistently from the original data this indicates systematic error is present.
10 microliters of the sample is then added and the assay absorption is measured at 340nm. If absorbance was above 1.5, samples were diluted.
Incorporation of assay controls included setting up a spectrophotomer and running the chart recorder with a full-scale deflection before the start of the assay. The set recorder had a corresponding value of 1 for the change in the absorbance. Therefore, prior testing was done to observe whether a change occurred in the readings. This helped to indicate that the results were valid, as they could have been affected by a fault during the setting up of the spectrophotometer. On the other hand this was considered as one of the controls for the experiment. Nevertheless, a new cuvette had to be used for each assay.
According to the scientific method, I should create a new experiment by making another hypothesis.
The absorbance is measured using a Plate reader and a Standard curve is generated. Also, the different types of pipetting techniques are assessed in this Assay.
Amylase concentration, the equation is y= 0.0382x - 1.0162. The R2 value is 0.92732. Serial dilutions were made using individual saliva, with measurements of 10x, 100x, and 1000x. From these dilution factors, the closest number to the absorbance curve was 1.4829ug/mL, which is my amylase concentration in saliva. During lab 6, the amylase saliva DNA was put into a PCR chain reaction and then in lab 7 it was put into a gel electrophoresis.
concentration, record the absorbance readings at a fixed wavelength, and plot the absorbance vs. concentration data. The wavelength of 520 nm was selected for experiment Part
The purpose of the experiment was to see if the object was alive. The hypothesis was if the object was alive it would grow in water. After adding water the data recorded indicated that the object had signs of life. The object turned from brown to a green the object grew and became softer. A blunder error could have been made in the process. The object could have been flooded with water because of an incorrect measurement. The hypothesis was supported by the data indicated that the object was alive.
The same solution of 0.5 ml BSA was then added from test tube 1 to the test tube 2 after being properly mixed, and from test tube 2 the solution was being added to test tube 3, and so forth all the way up to test tube 5, with the same exact procedure. From the last tube, we then disposed the 0.5 ml solution. After above procedures, we now labeled another test tube “blank”; 0.5 ml blank distilled water was purred into the tube with the serial dilution of 1:10. We also had a tube C labeled “unknown” with the same 0.5 ml of solution. And after adding 5ml of Coomassie Blue to each tube (1-5) and to the blank, the result of absorbance was read at 595 nm.
4. What is the most significant finding in the article? Summarize this clearly in several sentences.