A child was born without a working gene for an enzyme that the immune system needs. In 1990 her cells were repaired with a normal gene, and this was the first gene therapy. This lesson follows that treatment, and then shows how PCR and ELISA find a disease early.
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Repairing a gene
Some diseases are inherited. A child may be born with a gene that does not work. Could a doctor give the body a correct copy of the gene?
Gene therapy is the attempt to do this. It is a collection of methods that allow the correction of a gene defect that has been diagnosed in a child or an embryo. A normal gene is delivered into the cells and tissues of the person. It takes over the function of the non-functional gene and compensates for it.
The first gene therapy
The first clinical gene therapy was given in 1990 to a four-year-old girl with ADA deficiency. ADA is the enzyme adenosine deaminase. It is crucial for the immune system to function. The disorder is caused by the deletion of the gene for adenosine deaminase.
There were other treatments. In some children, bone marrow transplantation can cure the deficiency. In others, enzyme replacement therapy is used, in which the working enzyme is given by injection. But neither is completely curative.
A flow chart of gene therapy for ADA deficiency in five boxes. Box 1: lymphocytes are taken from the blood of the patient. Box 2: they are grown in a culture outside the body. Box 3: a functional ADA cDNA, carried by a retroviral vector, is introduced into the lymphocytes. Box 4: the genetically engineered lymphocytes are returned to the patient. A last note says that these cells are not immortal, so the patient needs periodic infusions, and that a gene put into early embryonic cells could be a permanent cure.
The new treatment ran in steps.
Lymphocytes were taken from the blood of the patient.
They were grown in a culture outside the body.
A functional ADA cDNA was introduced into the lymphocytes. A retroviral vector carried it. (A cDNA is a DNA copy of an mRNA, so it holds the coding part of the gene.)
The genetically engineered lymphocytes were returned to the patient.
These cells are not immortal. So the patient needs periodic infusions of such cells. A permanent cure may come from another method. If the ADA gene, isolated from the marrow cells that make the enzyme, is put into cells at an early embryonic stage, the cure could be permanent.
Finding a disease early (molecular diagnosis)
This part is not in the NEET syllabus text, but NCERT teaches it, and A-Level and IB ask about PCR and ELISA. It is kept out of the NEET format paper of this course.
For good treatment, early diagnosis matters. Conventional methods, such as the analysis of serum and urine, cannot find a disease early. A germ is usually suspected only when the disease shows symptoms. By then, the number of germs is already very high.
PCR, the polymerase chain reaction, copies a small piece of nucleic acid again and again. It can detect a very low number of bacteria or viruses before there are symptoms. It is used to detect HIV in a person who is suspected of AIDS. It can also find mutations in genes in suspected cancer and many other genetic disorders. The steps of PCR are in the previous chapter.
A probe is a single stranded DNA or RNA with a radioactive tag. It hybridises, which means that it pairs, with its complementary DNA in a clone of cells. The place of the probe is shown on a photographic film, which is called autoradiography. A clone with a mutated gene does not appear on the film, because the probe is not complementary to the mutated gene.
ELISA, the enzyme linked immuno-sorbent assay, is based on the interaction of an antigen with an antibody. An infection can be found from the antigens of the germ (proteins and glycoproteins) or from the antibodies that the body made against it.
Four small drawings of one well of a plate, in a row. Step 1: an antigen of the pathogen is stuck to the bottom of the well. Step 2: serum of the patient is added, and its antibodies, shown as Y shapes, bind to the antigen. Step 3: after washing, an antibody with an attached enzyme is added, and it binds to the antibodies of the patient. Step 4: a colourless substance is added, the enzyme changes it, and the well turns yellow. A note says that a colour means that the antibody, and so the infection, is present.
The steps of the picture show a common form of ELISA. NCERT states only the principle.
Try it in Python
The program counts the cycles of PCR needed to reach a number of copies that can be detected.
The second program reads an ELISA plate. The serum is diluted by half in each well.
Practise with new numbers
Common mistakes
Saying that gene therapy is a cure for ever. The lymphocytes are not immortal, so the infusions are repeated.
Mixing up the gene and the enzyme. In ADA deficiency the gene for adenosine deaminase is deleted. Enzyme replacement therapy gives the enzyme.
Saying that the ADA gene was put straight into the girl. The lymphocytes were grown outside the body, changed, and returned.
Thinking that PCR finds antibodies. PCR copies nucleic acid. ELISA finds antigens or antibodies.
Expecting a probe to bind to a mutated gene. A mutated gene is not complementary to the probe, so it does not appear on the film.
Remember these
Gene therapy delivers a normal gene to take over from a non-functional one. First case: 1990, ADA deficiency, a girl of four.
Steps: lymphocytes taken, grown outside the body, ADA cDNA put in by a retroviral vector, returned. Periodic infusions are needed.
A permanent cure may come from putting the gene into cells at an early embryonic stage.
PCR finds a low number of germs by copying their nucleic acid. ELISA uses antigen and antibody interaction.
Check yourself
Try each one on your own first. If you are stuck, open a hint. Then check with Show the answer.
What is gene therapy?
What is ADA deficiency, and what causes it?
Put in order: (a) return the cells to the patient; (b) take lymphocytes from the blood; (c) put in the ADA cDNA; (d) grow the cells outside the body.
Why does the patient need the infusion again and again?
How can PCR detect a virus before there are symptoms?