Enzyme-Linked Immunosorbent Assay (ELISA) — MCQs
All of the following forces are involved in antigen-antibody reactions, except:
HBsAg is based on which principle
What condition is known to cause a false positive ELISA for HIV?
Screening test for HIV infection in a patient prior to the development of antibodies (in window period):
Best method to diagnose HIV in an infant?
Which of the following is true about anti-CMV IgG antibodies?
Which one of the following enzymes is obtained from Thermus aquaticus bacterium that is heat stable and used in PCR at high temperature?
All of the following methods are used for the diagnosis of HIV infection in a 2 month old child, except –
The probe used in Western blot is:
Which of the following is a primarily RNA based technique?
Enzyme-Linked Immunosorbent Assay (ELISA) Indian Medical PG Practice Questions and MCQs
Question 1: All of the following forces are involved in antigen-antibody reactions, except:
- A. Electrostatic bond
- B. Hydrogen bond
- C. Covalent bond (Correct Answer)
- D. Van der Waals forces
Explanation: ***Covalent bond*** - **Covalent bonds** are strong, irreversible bonds that involve the sharing of electrons between atoms. - Antigen-antibody interactions are predominantly **non-covalent** and reversible, allowing for dynamic binding and release. *Vander Waal's forces* - **Van der Waals forces** are weak attractive forces that arise from temporary fluctuations in electron distribution, creating transient dipoles. - They are crucial in antigen-antibody binding, especially when the molecules are in **close proximity**, contributing to overall affinity. *Electrostatic bond* - **Electrostatic (ionic) bonds** occur between oppositely charged groups on the antigen and antibody surfaces. - These interactions are significant for **initial recognition** and overall binding stability, particularly at appropriate pH levels. *Hydrogen bond* - **Hydrogen bonds** form between a hydrogen atom covalently linked to an electronegative atom (like oxygen or nitrogen) and another electronegative atom. - They play a vital role in the **specificity and strength** of antigen-antibody interactions by providing numerous weak, directional contacts.
Question 2: HBsAg is based on which principle
- A. Chemiluminescence
- B. Immunofluorescence
- C. Immunochromatography assays
- D. ELISA (Correct Answer)
Explanation: ***ELISA*** - **Enzyme-linked immunosorbent assay (ELISA)** is a widely used laboratory test to detect and quantify antigens (like HBsAg) or antibodies in a sample. - It involves an enzyme-linked antibody that reacts with a substrate to produce a detectable signal, making it highly sensitive and specific for **HBsAg detection**. *Immunochromatography assays* - These are typically **rapid diagnostic tests (RDTs)** that provide quick qualitative results, often used for point-of-care testing. - While they can detect HBsAg, they generally have lower sensitivity and specificity compared to ELISA. *Chemiluminescence* - This is a detection method used in some immunoassays where a chemical reaction emits light, often providing higher sensitivity than colorimetric detection. - While it can be incorporated into HBsAg testing platforms, it is a *detection principle* rather than the primary assay principle like ELISA itself. *Immunofluorescence* - This technique uses **fluorescently labeled antibodies** to visualize antigens in cells or tissues under a fluorescence microscope [1]. - It is used for localization and identification of antigens, but not typically the primary method for routine quantitative HBsAg serology [1]. **References:** [1] Cross SS. Underwood's Pathology: A Clinical Approach. 6th ed. (Basic Pathology) introduces the student to key general principles of pathology, both as a medical science and as a clinical activity with a vital role in patient care. Part 2 (Disease Mechanisms) provides fundamental knowledge about the cellular and molecular processes involved in diseases, providing the rationale for their treatment. Part 3 (Systematic Pathology) deals in detail with specific diseases, with emphasis on the clinically important aspects., pp. 259-260.
Question 3: What condition is known to cause a false positive ELISA for HIV?
- A. Multiple myeloma
- B. Lepromatous leprosy
- C. Systemic lupus erythematosus (SLE) (Correct Answer)
- D. None of the listed conditions
Explanation: ***Systemic lupus erythematosus (SLE)*** - Patients with **SLE** can produce a variety of **autoantibodies**, some of which may cross-react with components of the HIV ELISA assay, leading to a false positive result [1]. - The immune dysregulation in SLE can cause **non-specific B-cell activation** and antibody production that confounds serological tests [1]. *Multiple myeloma* - This condition involves the uncontrolled proliferation of **plasma cells** and the overproduction of a single type of **monoclonal immunoglobulin** (M-protein). - While it can lead to various immune dysfunctions, it is not typically associated with false positive HIV ELISA results. *Lepromatous leprosy* - This severe form of leprosy is characterized by a **weak cell-mediated immune response** and high bacterial load, leading to widespread infection and hypergammaglobulinemia. - Though it involves substantial immune system changes, it is not a common cause of false positive HIV ELISA results. *None of the listed conditions* - This option is incorrect because **Systemic lupus erythematosus (SLE)** is a well-documented cause of false positive HIV ELISA results due to its complex autoimmune pathology [1].
Question 4: Screening test for HIV infection in a patient prior to the development of antibodies (in window period):
- A. Western blot
- B. p24 antigen (Correct Answer)
- C. ELISA
- D. All of the options
Explanation: ***p24 antigen*** - The **p24 antigen** test detects a structural protein of HIV, which becomes detectable before the development of antibodies (during the **window period**). - This test is crucial for early detection of HIV infection, especially when antibody tests might still be negative due to the time lag in immune response. *Western blot* - The **Western blot** is a confirmatory test for HIV, detecting specific antibodies to various HIV proteins. - It becomes positive only after the development of antibodies, making it unsuitable for detecting infection during the **window period**. *ELISA* - **ELISA** (Enzyme-Linked Immunosorbent Assay) is a common screening test for HIV, primarily detecting **HIV antibodies**. - Like Western blot, it relies on antibody presence and thus will be negative during the **window period**. *All of the options* - This option is incorrect because both **Western blot** and **ELISA** detect antibodies, making them unsuitable for screening during the **window period**. - Only the **p24 antigen** test can detect HIV infection before antibody seroconversion.
Question 5: Best method to diagnose HIV in an infant?
- A. ELISA
- B. PCR (Correct Answer)
- C. Western blot
- D. All of the options
Explanation: ***PCR*** - **Polymerase Chain Reaction (PCR)** detects **HIV nucleic acids** (DNA or RNA) directly, which is crucial for infants because maternal antibodies can persist for up to 18 months, interfering with antibody-based tests. - PCR allows for early diagnosis, often within the first few weeks or months of life, facilitating timely intervention. *ELISA* - **Enzyme-linked immunosorbent assay (ELISA)** detects HIV antibodies. - In infants, ELISA can be misleading due to the presence of **maternal HIV antibodies** transferred across the placenta, making it unreliable for diagnosing active infection. *Western blot* - **Western blot** is used to confirm positive ELISA results in adults by detecting specific HIV proteins. - Like ELISA, it relies on the detection of **antibodies** and is therefore not reliable in infants due to maternally transmitted antibodies. *All of the options* - This option is incorrect because **ELISA** and **Western blot** are antibody-based tests that are unreliable in infants due to the presence of **maternal antibodies**. - Only **PCR** directly detects the virus itself, making it the preferred diagnostic method in this age group.
Question 6: Which of the following is true about anti-CMV IgG antibodies?
- A. IgG avidity assay helps in differentiating past and primary infection (Correct Answer)
- B. Denotes latent CMV infection
- C. Denotes chronic CMV infection with immunity to other serotypes
- D. Indicates acute CMV infection
Explanation: ***IgG avidity assay helps in differentiating past and primary infection*** - **IgG avidity** measures the binding strength of IgG antibodies to their antigen. In a **primary infection**, IgG antibodies have low avidity. - As the immune response matures over several months, the avidity of IgG antibodies increases, indicating a **past infection**. *Denotes latent CMV infection* - While the presence of IgG antibodies indicates a past exposure and often a latent infection, it doesn't solely *denote* latency, as primary infection also involves IgG production. - **Latent CMV infection** specifically refers to the persistence of the virus in cells without active replication, which is usually confirmed by the presence of IgG antibodies but needs further contextual information like negative IgM and viral load. *Denotes chronic CMV infection with immunity to other serotypes* - CMV typically exists as one serotype, and IgG antibodies confer protection against *re-activation* of that specific virus, not immunity to "other serotypes." - **Chronic infection** usually implies ongoing active replication or persistent symptoms, which a positive IgG alone does not confirm. *Indicates acute CMV infection* - **Acute CMV infection** is primarily indicated by the presence of **IgM antibodies**, which appear early in the infection. - While IgG antibodies also rise during acute infection, their presence alone is not specific for an **acute phase** as they persist after the infection resolves.
Question 7: Which one of the following enzymes is obtained from Thermus aquaticus bacterium that is heat stable and used in PCR at high temperature?
- A. DNA gyrase
- B. DNA polymerase III
- C. Taq polymerase (Correct Answer)
- D. Endonuclease
Explanation: ***Taq polymerase*** - This **heat-stable DNA polymerase** is isolated from the thermophilic bacterium *Thermus aquaticus*. - Its ability to withstand high temperatures makes it ideal for the **polymerase chain reaction (PCR)**, where DNA denaturation steps occur at elevated temperatures. *DNA gyrase* - **DNA gyrase** is a type II topoisomerase that introduces negative supercoils into DNA, which is important for DNA replication and transcription. - It is not heat-stable and is not directly used for DNA amplification in PCR. *DNA polymerase III* - **DNA polymerase III** is the primary enzyme responsible for DNA replication in *E. coli* and other bacteria. - It rapidly synthesizes DNA but is **not heat-stable** and would denature at the temperatures required for PCR. *Endonuclease* - **Endonucleases** are enzymes that cleave phosphodiester bonds within a polynucleotide chain. - While essential for processes like DNA repair and restriction mapping, they are not primarily involved in and are not heat-stable for DNA synthesis in PCR.
Question 8: All of the following methods are used for the diagnosis of HIV infection in a 2 month old child, except –
- A. HIV ELISA (Correct Answer)
- B. Viral culture
- C. p24 antigen assay
- D. DNA–PCR
Explanation: ***Correct Answer: HIV ELISA*** (NOT used for definitive diagnosis in infants <18 months) - **HIV ELISA** detects **HIV antibodies**, which are maternally derived in infants born to HIV-positive mothers. - These maternal antibodies can persist up to **18 months**, leading to **false positive** results in uninfected infants, making ELISA **unreliable for definitive diagnosis** in infants younger than 18 months. - Therefore, ELISA is the correct answer to this "EXCEPT" question. *Incorrect: Viral culture* (CAN be used for diagnosis) - **Viral culture** involves growing the virus from the infant's blood, which is a highly **specific** method for detecting active HIV infection. - While sensitive, it is **expensive** and **time-consuming**, making it impractical for routine screening but still valid for diagnosis in infants. *Incorrect: p24 antigen assay* (CAN be used for diagnosis) - The **p24 antigen assay** directly detects a component of the **HIV capsid protein**, indicating active viral replication. - This method is effective for early diagnosis as it identifies the **virus itself**, not maternal antibodies, making it suitable for infant diagnosis. *Incorrect: DNA–PCR* (CAN be used for diagnosis) - **DNA–PCR (Polymerase Chain Reaction)** detects **proviral DNA** integrated into the host's cells, directly identifying the presence of HIV. - It is currently the **gold standard method** for early diagnosis in infants due to its high sensitivity and specificity in detecting actual infection.
Question 9: The probe used in Western blot is:
- A. Antibody (Correct Answer)
- B. mRNA
- C. DNA
- D. tRNA
Explanation: ***Antibody*** - In a **Western blot**, the primary probe used to detect specific proteins is typically an **antibody** that specifically binds to the target protein. - This antibody is often labeled (e.g., fluorescently or enzymatically) or recognized by a secondary, labeled antibody, allowing for visualization of the target. *mRNA* - **mRNA** (messenger RNA) is used as a probe in techniques like **Northern blotting** to detect specific RNA sequences, not proteins. - It carries genetic information from DNA to synthesize proteins but does not directly probe for protein presence in a Western blot. *DNA* - **DNA** is used as a probe in techniques such as **Southern blotting** to detect specific DNA sequences. - It is not used as a probe to identify proteins in a Western blot. *tRNA* - **tRNA** (transfer RNA) molecules are involved in **protein synthesis** by carrying specific amino acids to the ribosome. - They are not used as probes in any standard blotting technique, including Western blotting.
Question 10: Which of the following is a primarily RNA based technique?
- A. Sanger's technique
- B. Western blotting
- C. Next generation sequencing (Correct Answer)
- D. PCR
Explanation: ***Next generation sequencing*** - NGS, particularly **RNA-Seq (RNA sequencing)**, is the most advanced and comprehensive technique for studying RNA among the given options. - RNA-Seq directly sequences **RNA transcripts** to analyze gene expression, identify splice variants, detect novel transcripts, and quantify RNA abundance genome-wide. - While it involves converting RNA to cDNA for sequencing, **RNA is the primary starting material and target** of analysis, making it the most RNA-focused technique listed. - Superior to other methods for comprehensive transcriptome analysis. *Sanger's technique* - **Sanger sequencing** is a **DNA sequencing method** that determines the nucleotide sequence of DNA molecules. - Primarily designed for and used with DNA templates. - Not used for direct RNA analysis in routine practice. *Western blotting* - **Western blotting** detects and analyzes **proteins**, not nucleic acids. - Uses antibodies to identify specific proteins after electrophoretic separation. - Not related to RNA techniques. *PCR* - Standard **PCR amplifies DNA sequences** only. - While **RT-PCR** (reverse transcriptase PCR) starts with RNA, it immediately converts RNA to cDNA, and the actual amplification is DNA-based. - Less comprehensive for RNA analysis compared to RNA-Seq.
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