10 min readConcept sheet2 or 3-mark question on PCR and ELISA in early diagnosis; case-based questions; also a NEET favourite

Molecular diagnosis: PCR, probes and ELISA

Molecular diagnosis finds disease early. PCR amplifies tiny amounts of pathogen nucleic acid, radioactive probes find mutations, and ELISA detects antigens or antibodies.

Why molecular methodsConventional serum and urine tests cannot detect a disease early

PCR in diagnosisDetects very low amounts of a bacterium or virus by amplifying its nucleic acid; used for HIV and for gene mutations in suspected cancer

ProbeA single-stranded DNA or RNA tagged with a radioactive molecule, detected by autoradiography

ELISAEnzyme Linked Immuno-Sorbent Assay; based on antigen-antibody interaction

Why early diagnosis is hard

For good treatment, a disease must be found early. A pathogen is usually suspected only after it causes symptoms, and by then it is present in high numbers. Conventional methods such as serum and urine analysis cannot detect it early. Recombinant DNA technology, PCR and ELISA allow early diagnosis.

Uses of PCR in diagnosis

Detects a very low concentration of a bacterium or virus, before symptoms appear, by amplifying its nucleic acid.
Routinely used to detect HIV in suspected AIDS patients.
Detects mutations in genes in suspected cancer patients.
Identifies many other genetic disorders.

Finding a mutated gene with a radioactive probe

1
Make a probe: a single-stranded DNA or RNA tagged with a radioactive molecule.
2
Allow it to hybridise (base-pair) with its complementary DNA in a clone of cells.
3
Detect the bound probe by autoradiography: the radioactivity darkens a photographic film.
4
A clone with the normal gene shows up on the film.
5★
A clone with a mutated gene does not appear, because the probe is not complementary to it.

★ key step

ELISA

ELISA (Enzyme Linked Immuno-Sorbent Assay) works on the principle of antigen-antibody interaction. An infection can be detected by finding either the pathogen's antigens (proteins, glycoproteins) or the antibodies the body has made against it. In outline, a test well holds an antigen or antibody; the patient's sample is added; then an enzyme-linked antibody and a substrate are added. A colour change shows a positive result.

PCR vs ELISA

PCRDetects the pathogen's nucleic acid (DNA or RNA) by amplifying it. It can find a few copies, so it works very early.

ELISADetects proteins: the pathogen's antigens or the antibodies made against it, by antigen-antibody binding and a colour change.

Antibodies take time to form, so an antibody ELISA can miss a very early infection that PCR would find.

Choosing a test

Situation	Best test
A few HIV particles in the blood, before any symptoms	PCR
Checking whether a person has made antibodies against a virus	ELISA
Checking a clone of cells for a mutated gene	Radioactive probe and autoradiography
Looking for a gene mutation in a suspected cancer patient	PCR

In real life

During the COVID-19 pandemic, India set up RT-PCR labs across the country, and antibody surveys used ELISA kits. Blood banks screen donated blood for HIV and hepatitis with ELISA-type tests.

Exam angle

3 marks: “How do PCR and ELISA help in the early diagnosis of disease? State the principle of each.” Conventional tests miss early infection. PCR amplifies very small amounts of pathogen nucleic acid, so it finds HIV or gene mutations early. ELISA uses antigen-antibody interaction to detect the pathogen's antigens or the antibodies made against it.

2 marks: “How is a radioactive probe used to detect a mutated gene?” A radioactive single-stranded DNA or RNA probe hybridises with complementary DNA in a clone of cells; autoradiography shows bound probe. A clone with the mutated gene does not appear, as the probe is not complementary to it.

1 marks: “Expand ELISA.” Enzyme Linked Immuno-Sorbent Assay.

Remember it

PCR copies the code; ELISA spots the proteins.