# Must be 1500 word essay, on either topic 1 or 2. Harvard referencing with in text and reference list

i need to complete this assignment which is related to assay design course PHRM 4120: ELISA Calculations – Project Name: Date:
Calculating the results: First, complete the columns labeled OD1 and OD2 in the table below using the data from page #1. Then, calculate the mean OD for each sample. Write the mean value in the appropriate column. Construct the standard curve using the calculated values from the table. Plot the absorbance for the fT3 standards (mean) in the vertical axis (y) versus fT3 standard concentrations (T3 Levels pg/mL) in the horizontal axis (x) on a graph paper (you can use excel for this). Draw the best curve through the points. Attach the graph with your report. Read the concentration for each unknown sample from the curve. Record their value in the table below. Explain how the calculations of the unknown samples were performed under conclusions. Also under conclusions, provide a 3-5 sentences reflection of what you learned from this example. T3 levels Sample Identity OD 1 OD 2 Mean pg/ml Std1= 0.00 Std2= 1.20 Std3= 2.50 Std4= 5.00 Std5= 8.50 Std6= 18.00 Sample1= Sample2= Conclusions for Example #1: ___________________________________________________________________________________
Conclusions for Example #2: ___________________________________________________________________________________________
Questions about the assays in Examples #1 and #2: Compare and contrast the two ELISA procedures. Include at least 8 points of comparison. Why is never washed after the addition of the substrate in any ELISA assay? Of all the steps performed in ELISA, which is the most important and why? _____________________________________________________________________________
Example #3: ELISA to detect PCSK9/LDL receptor complexes Schematic representation of this assay: Plate setting and concentrations of standards: Answer the following questions about this assay: Compare and contrast this ELISA with the ELISA in example #2.

Reading of the plate at 450 and 540 nm: Answer the following questions about Example #3: Identify the following in this assay: Antigen: Antibodies: What type of ELISA is this assay? Explain your answer. Compare and contrast Examples #2 and #3. Use at least 8 points of comparison. Calculate the results: First, complete the columns labeled OD1 and OD2 for each wavelength in the table below using the data from page #12. Then, calculate the mean OD for each wavelength for each sample. Write the mean value in the appropriate columns. Subtract the mean OD at 540 nm from the mean OD at 450 nm (450 540; plate imperfection correction) for each sample and write the values in the appropriate column. Subtract standard 7 (blank or standard with concentration 0) from all the other samples (background subtraction) and write the values in the appropriate column. Construct the standard curve using the calculated double corrected values (*) from the table. Plot the absorbance for the complex standards in the vertical axis (y) versus complex standard concentrations (complex ng/mL) in the horizontal axis (x) on a graph paper (you can use excel for this). Draw the line through the points. Attach the graph with your report. Read the concentration for each unknown sample from the line. Record their value in the table below. Explain how the calculations of the unknown samples were performed under conclusions. Also under conclusions, provide a 3-5 sentences reflection of what you learned from this example. Sample Identity OD1 (450 nm) OD2 (450 nm) Mean OD1 (540 nm) OD2 (540 nm) Mean Correction (Subtract 540 from 450) Correction (Subtract Std 7 from all)* PCSK9/LDL receptor complexes (ng/ml) Std 1 = 20 Std 2 = 10 Std 3 = 5 Std 4 = 2.5 Std 5 = 1.25 Std 6 = 0.625 Std 7 = 0.000 0 Sample 1 = Sample 2 = Sample 3 = Sample 4 = Sample 5 = Conclusions about example #3: ___________________________________________________________________________________

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