ELISA

Question 1: In direct ELISA, which components are directly attached to the plate?
✓ Antigens
– Blocking agents
– Detection antibodies
– Capture antibodies

Question 2: Which component of sandwich ELISA is required in pairs?

✓ Antibodies
– Blocking agents
– Enzymes
– Antigens

Question 3: In a competitive ELISA, how would the signal measured by the detector react to an
increase of antigen in the sample?
✓ Decrease
– Match
– Not affected
– Increase

Question 4: Not just yet! We're actually missing one more step. Which step do we have to do next?
✓ Adding TMB substrate
– Incubating the plate
– Washing the plate
– Adding a horseradish peroxidase enzyme

Question 5: Genius! We're indeed missing one more step. Which step do we have to do next?
✓ Adding TMB substrate
– Incubating the plate
– Washing the plate
– Adding a horseradish peroxidase enzyme

Question 6: Marie: Why do we need to determine which cell lines produce the highest recombinant
FIXFactor nine?
✓ To reduce the cost of hemophilia treatment
– To increase the drug efficiency
– To hinder the spread of hemophilia
– To avoid cell contaminations

Question 7: Marie: The cell lines produce recombinant FIXfactor nine and other proteins. Which
biomolecule can specifically detect the presence of a certain protein, such as recombinant FIXfactor
nine, in a sample full of other proteins?
✓ Immunoglobulins
– Integrins
– Androgens
– Antigens

Question 8: Click View Image. What type of ELISA ELIZA is shown in the diagram?
✓ Indirect
– Competitive
– Sandwich
– Direct

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Question 9: In this coating step, the capture antibodies are immobilized on the surface of
polystyrene microplate wells. What type of force causes the passive adsorption of the captured
antibody due to the interaction between amino acid side chains on the antigen used for coating, and
the plastic surface?
✓ Hydrophobic
– Polar
– Van Der Waals
– Hydrophilic

Question 10: What are antibodies with high specificity that only detect one epitope called?
✓ Monoclonal
– Therapeutic
– Specific
– Polyclonal

Question 11: The basic blocking buffer contains 5% Bovine serum albumin (BSA) dissolved in PBS.
What is the purpose of adding a blocking buffer?
✓ To reduce ELISA background signal
– To minimize signal-to-noise ratio
– To create an optimal environment for hydrophilic interaction
– To prevent specific proteins from binding to the plate

Question 12: Tween 20 is used in our wash buffer. What type of compound is Tween 20?
✓ Surfactant
– Enzyme
– Substrate
– Reactant

Question 13: Why do we have to load the samples into a specific well?
✓ To achieve valid results
– To increase antibody affinity to samples
– To save reagents
– To avoid premature expiration

Question 14: What is the purpose of standard?

✓ To create standard curve
– To validate the false negative
– To avoid a false positive
– To ensure a high detection signal

Question 15: Quantitative ELISA includes a dilution series of known concentrations that is used to
create a standard curve. This standard curve allows the concentrations of antigens in a sample to
be quantified.

The first well contains 200 µL of standard with a concentration of 90 ng/mLnanograms per mililiter.
If we want to use a dilution with 1:2 dilution factor, how much sample should be transferred from the
first well to the second well that contains 100 µL diluent?
✓ 100 µL
– 150 µL
– 50 µL
– 200 µL

Question 16: We will perform serial dilution with 1:2 dilution factor. The concentration of the
standard in the first well is 90 ng/mL. The aliquot volume is 100 µL. The volume of diluent in well 2-

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8 is 100 µL each. What is the concentration of the standard in well 8? Take into account that we are
diluting the concentration to half in each step.
✓ 0.7 ng/µL
– 1.7 ng/µL
– 11.3 ng/µL
– 5 ng/µL

Question 17: A positive control is a sample known to give positive results for the given test. What is
the purpose of positive control?
✓ Verify that the negative results are valid
– Validate false positive results
– Always give positive results
– Give positive results even when the procedure is not optimized

Question 18: What is a group of samples where no response is expected called? This group
contains all buffers and reagents except the substance of interest.
✓ Negative control
– Test sample
– Positive sample
– Standard

Question 19: What is the purpose of agitating the ELISA plate?

✓ Increasing the rate of binding
– Optimizing the signal-to-noise ratio
– Reducing the washing time
– Prolonging the incubation time

Question 20: TMB (tetramethyl benzene) is a substrate for horseradish peroxidase (HRP). What is
the proper container for storing TMB?
✓ Light protected container
– Metal vial
– Glass beaker
– Eppendorf tube

Question 21: A stop solution is added to provide a fixed end point for the assay. What is the stop
solution for the ELISAELIZA substrate TMB in the presence of horseradish peroxidase (HRP)?
✓ Sulfuric acid
– Hydrochloric acid
– Sodium chloride
– Sodium hydroxide

Question 22: Click "Explore" to access the graphs on the PC screen.

A regression formula is obtained from the standard curve. This formula describes the relationship
between concentration and absorbance data. We can use this regression formula to calculate the
concentration of unknown samples. In this case, which regression type best describes our
absorbance data?
✓ Log-log
– Power
– Exponential
– Linear

Question 23: The equation of the standard curve is y = 0.9988x - 1.437why equals zero point nine
nine eight eight X minus one point four three seven. In ELISAELIZA we measured the absorbance
using a spectrophotometer. How can we calculate the amount of Factor nineIX (FIX) in each

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sample?
✓ Amount of FIX factor nine= (Absorbance + 1.437)/divided by0.9988
– Amount of FIXfactor nine = (0.9988 + Absorbance)/divided by1.437
– Amount of FIXfactor nine *times 0.9988 = Absorbance + 1.437
– Amount of FIXfactor nine = 0.9988 *times Absorbance - minus by 1.437

Question 24: From: Ms. Stauffer.

Hello, I need your help for troubleshooting. My ELISAELIZA result looks weird. Click 'View Image' to
see it.

All wells exhibit similar bright blue colors, even the negative control. What kind of problem do I
have?
✓ You have high background
– You have no color development
– You have low absorbance
– You have no optical reading

Question 25: From: Senor Guzman.

Hi! I have generated the standard curve for my ELISA experiment. Click 'View Image' to see it.
What do you think?
✓ Your standard curve is poor
– You're using the wrong regression
– You don't have enough samples
– Perfect, standard curve Senor!

Question 26: Alright! Marie has completed the remaining parts of the data analysis. The average
absorbance of each sample are as follows:

Human kidney cell lines (Test sample 1): 8.2

Human hepatic cell lines (Test sample 2): 5.7
Human pancreatic cell lines (Test sample 3): 3.6

Which cell lines produce the highest amount of recombinant FIXfactor nine?
✓ Human kidney cell lines
– Cannot be determined
– Human pancreatic cell lines
– Human hepatic cell lines

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