Subtopic: Fundamentals of Biomolecules, enzyme kinetics
Question Statement:
In ELISA, which of the following enzymes are conjugated to antibodies for
detection of the analyte?
P. Alkaline phosphatase
Q. Trypsinase
R. Horseradish peroxidase
S. Amylase
Options:
(A) P and R
(B) P and Q
(C) Q and S
(D) R and S
Official Answer Key:
A
Step-by-Step Solution & Explanation
### **Official Solution & Concept Analysis (GATE BT 2023 Q.22)**
**1. Governing Concept**:
- **Topic**: Immunological Techniques & Diagnostic Enzymology (ELISA)
- In Enzyme-Linked Immunosorbent Assay (ELISA), reporter enzymes conjugated to primary or secondary detection antibodies must have high turnover numbers, stable covalent conjugation, and easily assayable chromogenic/fluorogenic substrates.
- The two most universally utilized reporter enzymes are:
- **P. Alkaline phosphatase (ALP)** (hydrolyzes p-nitrophenyl phosphate, pNPP, yielding yellow p-nitrophenol).
- **R. Horseradish peroxidase (HRP)** (oxidizes substrates like TMB, OPD, ABTS in the presence of $\text{H}_2\text{O}_2$).
**2. Option Analysis**:
- **Option (A) [P and R] is CORRECT**: Alkaline phosphatase and horseradish peroxidase are the universal ELISA conjugates.
- **Option (B) is INCORRECT**: Trypsin is a digestive endopeptidase, not an ELISA reporter.
- **Option (C) is INCORRECT**: Neither trypsinase nor amylase are standard diagnostic conjugates.
- **Option (D) is INCORRECT**: Amylase is not used as an ELISA detection enzyme.
**3. Final Answer**:
- **Correct Choice**: **A**
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