ELISA and Western Blotting: Principles and Applications in Immunology

Question 1

What is the primary purpose of a standard curve in Western blotting?

Accepted Answer

Determining the concentration of the target protein in the sample

Answer

Identifying the molecular weight of the target protein

Answer

Determining the specificity of the primary antibody

Answer

Validating the reproducibility of the blotting technique

Question 2

Which is the primary advantage of Western blotting over ELISA assays?

Accepted Answer

Ability to detect multiple proteins simultaneously

Answer

Higher sensitivity

Answer

Quantitative analysis of protein expression

Answer

Simplicity and cost-effectiveness

Question 3

Which of the following steps is not involved in Western blotting?

Accepted Answer

Hybridization with a labeled secondary antibody

Answer

Protein separation by electrophoresis

Answer

Transfer of proteins to a nitrocellulose membrane

Answer

Incubation with primary antibody

Question 4

What is the primary role of blocking buffer in Western blotting?

Accepted Answer

Preventing non-specific binding of antibodies

Answer

Enhancing the sensitivity of the assay

Answer

Fixing the proteins on the membrane

Answer

Reducing background noise

Question 5

What is the significance of enzyme conjugates in ELISA assays?

Accepted Answer

Enabling colorimetric or fluorescent detection

Answer

Enhancing the specificity of the antibody-antigen reaction

Answer

Increasing the sensitivity of the assay

Answer

Preventing non-specific binding

Question 6

What is the purpose of chemiluminescence substrates in Western blotting?

Accepted Answer

Producing a light signal for protein detection

Answer

Blocking non-specific binding of antibodies

Answer

Enhancing the sensitivity of the assay

Answer

Fixing the proteins on the membrane

Question 7

In Western blotting, which of the following is the correct order for these essential steps: cell lysis, protein extraction, and protein quantification?

Accepted Answer

Cell lysis, protein extraction, protein quantification

Answer

Protein quantification, protein extraction, cell lysis

Answer

Cell lysis, protein quantification, protein extraction

Question 8

What is the primary function of the blocking step in a Western blot procedure?

Accepted Answer

To prevent non-specific binding of antibodies to the membrane

Answer

To increase the sensitivity of the assay

Answer

To enhance the binding of the secondary antibody

Answer

To remove excess primary antibody

Question 9

For the antigen-antibody reaction in ELISA, what is the optimal pH range?

Accepted Answer

6.8-7.2

Answer

4.5-5.5

Answer

8.0-8.5

Answer

9.0-9.5

Question 10

In Western blotting, which enzyme conjugate is commonly used?

Accepted Answer

Horseradish peroxidase (HRP)

Answer

Alkaline phosphatase (AP)

Answer

Luciferase

Answer

beta-Galactosidase

Question 11

What is the purpose of the substrate in an ELISA assay?

Accepted Answer

To produce a colorimetric signal

Answer

To amplify the antigen-antibody reaction

Answer

To block non-specific binding

Answer

To enhance the binding of the primary antibody

Question 12

Which of the following is NOT a potential application of Western blotting?

Accepted Answer

Gene sequencing

Answer

Protein expression analysis

Answer

Post-translational modification detection

Answer

Drug target identification

Question 13

ELISA utilizes which principle for its operation?

Accepted Answer

Antigen-antibody interaction

Answer

DNA amplification

Answer

Cell surface marker analysis

Answer

Nucleic acid hybridization

Question 14

Western blotting is commonly employed for:

Accepted Answer

Detection and identification of specific proteins

Answer

Quantification of gene expression levels

Answer

Sequencing of DNA fragments

Answer

Identification of bacterial species

Question 15

In Western blotting, chemiluminescence substrates like SuperSignal are used for:

Accepted Answer

Producing light signals for detection

Answer

Enhancing protein separation

Answer

Blocking non-specific antibody binding

Answer

Visualizing DNA fragments

Question 16

The blocking step in Western blotting primarily aims to:

Accepted Answer

Prevent non-specific antibody binding

Answer

Increase signal intensity

Answer

Enhance protein separation

Answer

Detect the target protein

Question 17

In Western blotting, protein separation is achieved through:

Accepted Answer

Gel electrophoresis

Answer

Chromatography

Answer

Polymerase Chain Reaction (PCR)

Answer

Flow cytometry

Question 18

The purpose of a positive control in Western blotting is to:

Accepted Answer

Confirm the specificity of antibodies

Answer

Determine optimal antibody concentration

Answer

Calibrate the chemiluminescence detector

Answer

Detect the presence of the target protein

Question 19

The distinction between primary and secondary antibodies lies in their specificity:

Accepted Answer

Primary antibodies bind to the target protein, while secondary antibodies bind to the constant region of the primary antibody.

Answer

Primary antibodies bind to the target organism, while secondary antibodies bind to a different organism.

Answer

Primary antibodies are used in direct ELISA, while secondary antibodies are used in indirect ELISA.

Answer

There is no distinction between primary and secondary antibodies.

Question 20

In an indirect ELISA, after the antigen is bound to the well, what is the next step?

Accepted Answer

A primary antibody specific to the antigen is added, followed by a secondary antibody conjugated to an enzyme.

Answer

A secondary antibody specific to the antigen is added, followed by a primary antibody conjugated to an enzyme.

Answer

A substrate is added to react with the enzyme, producing a detectable signal.

Answer

An enzyme-linked antigen is added to bind to the primary antibody.

Question 21

Which of the following is NOT a typical application of ELISA?

Accepted Answer

Determining the size of a protein.

Answer

Screening for infectious diseases.

Answer

Measuring the concentration of a specific protein in a sample.

Answer

Detecting the presence of antibodies in a sample.

Question 22

In Western blotting, proteins are separated based on what characteristic?

Accepted Answer

Molecular weight.

Answer

Isoelectric point.

Answer

Charge.

Answer

Hydrophobicity.

Question 23

A researcher wants to determine if a patient's serum contains antibodies against a specific viral protein. Which technique is most suitable for this task?

Accepted Answer

Indirect ELISA.

Answer

Western blotting.

Answer

Sandwich ELISA.

Answer

Direct ELISA.

Question 24

What is a crucial step in both ELISA and Western blotting that helps prevent non-specific binding?

Accepted Answer

Blocking non-specific binding sites.

Answer

Adding a substrate to generate a detectable signal.

Answer

Performing gel electrophoresis.

Answer

Using radioactive labels.

Question 25

In Western blotting, a strong signal indicates what?

Accepted Answer

A high concentration of the target protein in the sample.

Answer

A weak binding affinity between the antibody and the antigen.

Answer

A high concentration of antibodies in the sample.

Answer

A successful separation of proteins based on their size.

Question 26

Which type of ELISA is commonly used to detect the presence of a specific antigen in a sample?

Accepted Answer

Sandwich ELISA.

Answer

Direct ELISA.

Answer

Indirect ELISA.

Answer

Competitive ELISA.

Question 27

What is the main purpose of using a secondary antibody in ELISA and Western blotting?

Accepted Answer

To amplify the signal and improve detection sensitivity.

Answer

To separate proteins based on molecular weight.

Answer

To block non-specific binding sites.

Answer

To bind directly to the antigen.

Question 28

A researcher observes multiple bands on a Western blot, suggesting the presence of different protein isoforms. What is the most likely explanation for this result?

Accepted Answer

The protein has undergone post-translational modifications.

Answer

The antibody used is non-specific.

Answer

The sample was contaminated.

Answer

The protein was not properly separated during gel electrophoresis.