At first, we prepared a 5 % fresh milk, that is for 30 mL PBS we added 1.5 g of powdered milk. Then we removed membrane from the “sandwich” and placed it on the cutting board, and folded plastic wrap around membrane. The size of the target protein is 100 kDa, so we cut it on 150 kDa and below 75 kDa. After removing plastic wrap and labeling the membrane, we put it in the 5% milk and placed it on the bench-top for 30 minutes agitation at room temperature. Powdered milk also contains many proteins, which coat the surface of the membrane. The coating of the membrane surface block our antibody from binding non-specifically to other proteins. After 30 minutes of agitation, membrane was rinsed 3 times in 1X PBST with gentle shaking, using a generous amount of buffer. Additionally, excess milk was removed by washing it 3 times …show more content…
We washed off the primary antibody from a membrane at the same manner as we washed off the milk, and then incubated the blot for an hour at room temperature with the secondary antibody GAR680, from this point a membrane was shielded from the light until it has been imaged. The secondary antibody specifically recognizes the primary antibody. GAR680 is Goat Anti-Rabbit immunoglobin antibody. This is an antibody raised in goats that recognizes rabbit immunoglobin. Excess secondary antibody is removed by washing in PBST buffer as was done for the primary antibody, the final wash with 1X PBS to remove residual detergent Tween 20. The membrane was ready for scan and reading on Li-Cor machine. Specially designated tray for fluorescent antibodies was cleaned up with deionized water and kimwipes, then only the membrane was placed on it and entered the Li-Cor machine. The membrane position was from the first to the 10th well, from left to the right, the label was on the left side. The membrane reading was made on 700 nm
That familiar fizzing you hear when you drop an Alka Seltzer tablet into a glass of water is the result of a chemical reaction, and chemical reactions are extremely prevalent when it comes to what living things do to carry out life processes. In addition, environmental conditions can alter the results of chemical reactions, and in this lab, we will be answering the
Forensic Science Introduction: Someone in a restaurant has suddenly fallen ill and a mystery powder has been discovered with the victim. As the chief investigator, your duty is to identify the mystery substance through a lab. In this lab, it will consist of five known compounds and one unknown compound. Your job is to distinguish which one out of the five substances is the mystery powder. To figure out the mystery matter you will have to compare their physical and chemical properties and match them with the appropriate compound.
The affects of pH, temperature, and salt concentration on the enzyme lactase were all expected to have an effect on enzymatic activity, compared to an untreated 25oC control. The reactions incubated at 37oC were hypothesized to increase the enzymatic activity, because it is normal human body temperature. This hypothesis was supported by the results. The reaction incubated to 60oC was expected to decrease the enzymatic activity, because it is much higher than normal body temperature, however this hypothesis was not supported. When incubated to 0oC, the reaction rate was hypothesized to decrease, and according to the results the hypothesis was supported. Both in low and high pH, the reaction rate was hypothesized to decrease, which was also supported by the results. Lastly, the reaction rate was hypothesized to decrease in a higher salt concentration, which was also supported by the results.
One of the most primitive actions known is the consumption of lactose, (milk), from the mother after birth. Mammals have an innate predisposition towards this consumption, as it is their main source of energy. Most mammals lose the ability to digest lactose shortly after their birth. The ability to digest lactose is determined by the presence of an enzyme called lactase, which is found in the lining of the small intestine. An enzyme is a small molecule or group of molecules that act as a catalyst (catalyst being defined as a molecule that binds to the original reactant and lowers the amount of energy needed to break apart the original molecule to obtain energy) in breaking apart the lactose molecule. In mammals, the lactase enzyme is present
Mader, S. S. (2010). Metabolism: Energy and Enzymes. In K. G. Lyle-Ippolito, & A. T. Storfer (Ed.), Inquiry into life (13th ed., pp. 105-107). Princeton, N.J: McGraw Hill.
The radioallergosorbent test (RAST) is an RIA test that allows for the detection and quantification of IgE antibodies. During the test, a possible allergen is bound to insoluble material and a sample of the patient’s blood is added. If the blood sample contains antibodies specific to that allergen, the antibodies will bind to that allergen. Anti-human radiolabeled IgE antibodies are then added and bind only to the antibodies already bound to the insoluble material. The whole sample is then wThe unbound anti-human antibodies are washed away and the level of radioactivity is proportional to the amount of human IgE present in the blood sample.
Several researchers have evaluated the specific antibody response that is responsible for HAR. An in vitro kinetic experiment combined rat endothelial cells with primate serum and then measured bound human and monkey antibodies, number of lysed cells, and C complement activity (Azimzadeh et al., 1996). The results showed that IgM antibodies were produced rapidly in the earliest stage, after which a large number of IgG antibodies were produced. Components of the C cascade were present on the endothelial cells. Th...
Solid A was identified to be sodium chloride, solid B was identified to be sucrose, and Solid C was identified to be corn starch. Within the Information Chart – Mystery White Solid Lab there are results that distinguishes itself from the other 4 experimental results within each test. Such as: the high conductivity and high melting point of sodium chloride, and the iodine reaction of corn starch. Solid A is an ionic compound due to its high melting point and high electrical conductivity (7), within the Information Chart – Mystery White Solid Lab there is only one ionic compound which is sodium chloride, with the test results of Solid A, it can be concluded that is a sodium chloride. Solid B was identified as sucrose due to its low electrical
Anne Zhang 3/6/14 BSGE 7-1 Lab Report Problem Paragraph 1 Question: What is the effect of temperature on the dissolving time of an Alka-Seltzer? Alka-Seltzer is made up of baking soda, aspirin, and citric acid which gives the tablet the fizz when dropped in any temperature water. “Alka-Seltzer is a medication that works as a pain reliever and an antacid.
In this experiment, we determined the isotonic and hemolytic molar concentrations of non-penetrating moles for sheep red blood cells and measured the absorbance levels from each concentration. The results concluded that as the concentration increased the absorbance reading increased as well. A higher absorbance signifies higher amounts of intact RBCs. The isotonic molar concentration for NaCl and glucose is 0.3 M. The hemolysis molar concentration for NaCl and glucose is 0.05 M. Adding red blood cells to an isotonic solution, there will be no isotonic pressure and no net movement. The isotonic solution leaves the red blood cells intact. RBC contain hemoglobin which absorbs light, hemoglobin falls to the bottom of the tube and no light is absorbed. Determining the isotonic concentration of NaCl and glucose by finding the lowest molar concentration. In contrast to isotonic molar concentration, hemolysis can be determined by finding the
In our Biology Lab we did a laboratory experiment on fermentation, alcohol fermentation to be exact. Alcohol fermentation is a type of fermentation that produces the alcohol ethanol and CO2. In the experiment we estimated the rate of alcohol fermentation by measuring the rate of CO2 production. Both glycolysis and fermentation consist of a series of chemical reactions, each of which is catalyzed by a specific enzyme. Two of the tables substituted some of the solution glucose for two different types of solutions. They are as followed, Table #5 substituted glucose for sucrose and Table #6 substituted the glucose for pH4. The equation for alcohol fermentation consists of 6 Carbons 12 Hydrogens 6 Oxygen to produce 2 pyruvates plus 2 ATP then finally the final reaction will be 2 CO2 plus Ethanol. In the class our controlled numbers were at Table #1; their table had 15 mL Glucose, 10 mL RO water, and 10 mL of yeast which then they placed in an incubator at 37 degrees Celsius. We each then measured our own table’s fermentation flasks every 15 mins for an hour to compare to Table #1’s controlled numbers. At
According to the graph on amylase activity at various enzyme concentration (graph 1), the increase of enzyme dilution results in a slower decrease of amylose percentage. Looking at the graph, the amylose percentage decreases at a fast rate with the undiluted enzyme. However, the enzyme dilution with a concentration of 1:3 decreased at a slow rate over time. Additionally, the higher the enzyme dilution, the higher the amylose percentage. For example, in the graph it can be seen that the enzyme dilution with a 1:9 concentration increased over time. However, there is a drastic increase after four minutes, but this is most likely a result of the error that was encountered during the experiment. The undiluted enzyme and the enzyme dilution had a low amylose percentage because there was high enzyme activity. Also, there was an increase in amylose percentage with the enzyme dilution with a 1: 9 concentrations because there was low enzyme activity.
= Before conducting the experiment I would conduct a simple test for the protein by placing a sample of the albumen into a test tube and add biurett reagent. This contains copper (II) sulphate and sodium hydroxide.
4. Put milk samples into the beaker for about five and a half minutes and take samples out after time is up. 5. With the warm samples, open the pouch containing the gel cassette and remove the cassette.