Test 3 - d. How was this ELISA reaction set up? Draw a...

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1. Why do we typically prefer to use monoclonal antibodies? What is the major drawback of monoclonal antibodies? How are monoclonal antibodies made? (Keep it general – no more detail than was given in lecture.) 2. You have just prepared a monoclonal antibody that has a very high affinity for your favorite protein. Your protein has three different subunits (10 kDa, 25 kDa, and 55 kDa), and you need to know which subunit binds your new antibody. What is a simple way to determine this? 3. You are performing an ELISA reaction to measure estradiol levels in unknown samples. You also perform this reaction on a series of known standards. Your results from the standards are shown in the graph below: 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 0 20 40 60 80 Estradiol concentration (pg/mL) Absorbance a. Why do we need to include the known standards? b. How will your experimental protocol differ between the standards and your unknown? c. What is the concentration of your unknown if your absorbance readings average 0.44?
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Unformatted text preview: d. How was this ELISA reaction set up? Draw a diagram of a well and the components in it. (There are several possibilities try to keep it simple!) 1. Western blotting is the electrophoretic transfer of proteins from a gel to a filter. a. How do we set up the electrodes for the transfer with respect to the gel and the membrane? (Diagram is probably the easiest here.) b. What type of forces bind the protein to the membrane? c. What ingredient in the transfer buffer helps to ensure that these forces (from part b) are as strong as possible? How does this work? 2. Pregnancy tests detect the hormone hCG (human chorionic gonadotropin), which is produced by the embryo. On the following templates (not shown, but it is just a basic test) , label the reagents present at each zone, draw a simple diagram showing the antibodies and antigens (include labels!), and draw where you would expect to see color in the positive and negative tests....
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