What Is Molecular Testing? How Is It Done? Uses in Cancer and Other Diseases

What Is Molecular Testing?

Molecular tests examine the structure and characteristics of biological materials, such as DNA, RNA, proteins or fats, by taking samples of these materials.

These tests are used in diagnosing diseases, planning treatment and follow-up. For example, molecular diagnostic tests may look for influenza-virus or coronavirus DNA to establish whether someone has flu or COVID-19. Similarly, cancer-cell DNA can be examined to identify a cancer subtype, cancer-causing mutations or eligibility for targeted drugs. Molecular tests are also frequently used in allergy and immunology.

Molecular diagnostic tests are important in medicine because they provide highly sensitive and accurate results.

Are Molecular and Genetic Tests the Same?

Molecular tests are often called “genetic tests”. However, they are not identical. They may serve similar purposes but provide different information by examining different biological materials. Molecular tests can, for example, examine biomarkers such as proteins and fat molecules. Genetic tests examine a person’s genetic makeup, DNA and RNA.

Types of Molecular Tests

Molecular tests use many techniques to detect molecules in samples and make them more visible. Samples generally come from saliva, blood or tissue biopsy material.

Molecular tests offer greater certainty in many settings. We use them, for example, to identify genetic changes specific to a cancer type and select more targeted treatments. They look for DNA or RNA segments associated with particular conditions and provide definitive results when performed correctly. Patients with a provisional diagnosis may later receive a definitive diagnosis through molecular testing. 

Many molecular testing techniques are available:

• Polymerase chain reaction (PCR): This amplifies selected DNA regions in biological samples. PCR is fast and sensitive and forms the basis of many molecular tests.

• Next Generation Sequencing (NGS): This determines the complete nucleotide sequence of a DNA sample. It is important for research such as identifying genetic mutations and reconstructing evolutionary trees from biological samples.

• Southern blotting: This separates DNA by electrophoresis and then detects a specific region. It is used to identify genetic mutations and chromosomal abnormalities.

• Western blotting: This separates proteins by electrophoresis and detects bands belonging to a specific protein. It is used to identify protein expression levels or modifications.

• Fluorescence in situ hybridization (FISH): This identifies the presence or location of a gene in cells. FISH uses fluorescently labeled nucleotides matching the DNA sequence of the gene for which it is designed. They bind to the target gene’s DNA and are examined under a special light-emitting microscope, allowing the gene’s presence or location to be determined.

• Mass spectrometry: This determines molecular structure by measuring the weights of atoms in a substance, identifying the proportions of different atoms and providing structural information.

• NMR (nuclear magnetic resonance) spectroscopy: This determines molecular structure by measuring the magnetic properties of atoms. It identifies the positions of different atoms and provides information about the molecule’s structure.

For each technique, specialists take a bodily sample and make its molecules more visible. A specialist then examines it under a microscope or with specialized biomedical equipment and reports the findings.

Why Are Molecular Tests Ordered?

Specialists use molecular tests to gain more information for diagnosis and predict outcomes. They serve many purposes, including:

• Assessing risk for various diseases
• Confirming infections undetectable by conventional laboratory techniques
• Genetic counseling for people who are pregnant or planning pregnancy
• Supporting treatment decisions
• Assessing how well someone may respond to a drug
• Diagnosing subtypes of cancers such as breast, lung and colon cancer

A Common Molecular Test in Cancer: HER2 FISH

The HER2 FISH test is a molecular technique for detecting overproduction of the HER2 gene in breast cancer cells. HER2 is a protein-producing gene found in breast tissue. Normally, HER2 gene production is regulated and balanced. Sometimes, however, HER2 gene overproduction occurs, causing rapid cell growth and proliferation, which can trigger breast cancer.

HER2 FISH uses fluorescent nucleotides matching the target gene’s DNA sequence. They bind to its DNA and are examined with a special light-emitting microscope. This detects overproduction (overexpression) of the HER2 gene product, the HER2 protein. HER2 FISH is used for diagnosis and treatment guidance. For example, patients with overproduction of the HER2 gene product benefit more from HER2-targeted precision drugs in their treatment.

What Is Needed for Molecular Testing?

Each test requires a sample from the person. COVID-19 tests, for example, look for coronavirus-specific antigens in nasal and respiratory secretions. Antigens are typically present on viral surfaces.

Possible samples include:

• A tumor fragment obtained by biopsy
• Blood
• Body fluids such as respiratory secretions
• Amniotic fluid

2 Main Classes of Molecular Tests

Molecular testing is developing rapidly, with growing numbers of tests. COVID-19 PCR tests, for example, are a type rapidly developed by scientists to address the COVID-19 pandemic.

In general, molecular tests fall into two broad categories with many different subtypes:

1. Genome sequencing and testing
2. Genetic biomarkers

Genome Sequencing and Testing

Genome sequencing involves analyzing all or part of a cell's genetic material to look for particular genes or gene changes. Genes are made of DNA and provide instructions for the cell to make proteins. Genome sequencing can also assess whether a person carries a gene for a disease. It is a common type of test performed during pregnancy or when planning to become pregnant.

Another use of genome sequencing is to help predict a person's response to certain medicines. For example, carriers of particular genes may not respond well to some medicines or may have a higher risk of developing certain allergic reactions.

Advantages and Disadvantages of Molecular Tests

These types of tests offer many advantages, including the following:

• Prediction: If a person carries genes for certain harmful or potentially life-threatening diseases, this can help with family planning.
• Treatment decisions: A person at higher risk of developing certain diseases can make long-term plans based on this information or change their lifestyle to reduce their risk. Testing for various cancer-associated genetic variants can guide breast cancer screening.
• Other uses: Some people may seek molecular testing for inherited traits to learn about potential hereditary diseases in their ancestry or because they are curious about their genetic heritage.

There are also some disadvantages:

• Reliability: Test reliability can vary, particularly for inherited genetic testing. Tests that identify genetic risk factors do not guarantee that a person will develop a disease and may cause unnecessary anxiety.
• Decision-making: Making decisions based on genetic information can be difficult. For example, a person may have to decide whether to end a pregnancy after learning that their child has a 50% chance of carrying a life-threatening disease.
• Accessibility: Access to molecular tests may be limited because they can be expensive and may not be covered by insurance.
• Privacy: Choosing companies that are not accredited and monitored for genetic testing can jeopardize a person's genetic privacy.

Genetic Markers

Another type of genetic test looks for genetic biomarkers of infection or other diseases. Rather than sequencing a person's genome, these tests look for embedded genetic signals that have a strong association with the suspected disease.

The COVID-19 PCR test is one example of this type. This test looks for genetic material carried by the virus that causes COVID-19.

In oncology, genetic testing is also performed on certain types of cancer tumors. Different genetic signatures can provide information about the type of cancer a person has and which treatment is more likely to be effective.

The main benefit of genetic biomarker testing is its high likelihood of accuracy. In some cases, these tests can confirm a diagnosis. They can also help specialists make well-informed medical decisions or better guide treatment.

What Are the Disadvantages of Genetic Markers?

• Accessibility: Molecular tests are generally more expensive and less accessible. For example, cancer genetic tests cost more and take longer to produce results. Similarly, the COVID-19 PCR test is more costly and takes longer to produce results than home antigen tests. In addition, not every patient may be suitable for every molecular test.
• Reliability: Molecular tests for diseases and infections are generally accurate, but no test offers 100% reliability.
• Sample problems: The samples obtained may be insufficient for testing, and the process may be hindered, as when a biopsy does not yield enough tissue.

Molecular Tests and COVID-19

Polymerase chain reaction (PCR) tests are among the most accurate and widely used molecular tests for detecting COVID-19. They work by looking for diagnostic genetic material specific to the virus and then amplifying this small piece of viral DNA for laboratory confirmation. It generally takes a few days to obtain a result from these tests.

What Is the Difference Between Antigen and Molecular Tests for COVID-19?

Antigen tests look for viral proteins. These proteins are easier to detect, and the tests can quickly give a positive or negative result. PCR tests look for viral genetic material and then amplify it so that diagnostic technology can confirm the presence of the virus. These tests are more sensitive and complex. They can detect the virus before a person becomes symptomatic or infectious.

What Is Molecular Allergy Testing?

Molecular allergy testing is a test used to determine a person's sensitivity to a particular allergen (allergens are substances that can cause an allergic reaction in the human body). For example, after someone is bitten by an unidentified insect, molecular allergy testing can be used to determine what type of insect it was.

Molecular allergy tests are generally performed using samples taken from the blood or nose. For example, a sample of mucus from a person's nose can be taken and examined for allergens. Similarly, a person's blood sample can be taken and examined for allergens. By determining whether a person is sensitive to a particular allergen, these tests help identify the causes of allergy symptoms.

Frequently Asked Questions About Molecular Tests

How can molecular tests help with cancer treatment?

Molecular tests can help guide treatment for some cancer patients. However, molecular testing may not be useful in every situation, including the following:

• If a tissue biopsy cannot be obtained
• If the biopsy sample is insufficient

However, liquid biopsies (from blood or other fluids) have also begun to become more widely used in cancer when tissue samples are insufficient.

Sometimes, even if molecular tests point to a particular type of treatment, there is no guarantee that it will treat the cancer effectively.

What is Next Generation Sequencing (NGS)?

Next generation sequencing is a type of high-speed molecular testing that can sequence many different DNA segments at the same time. It is more sensitive and reliable than conventional techniques. Next generation sequencing can also detect smaller genetic variations and provides a more comprehensive view of a person's genome and genetic risk factors.

NGS has now become the most reliable and most widely used molecular test in cancer genetic testing.

How accurate are molecular tests?

The accuracy of molecular testing depends on the test. In general, these tests are accurate but not perfect. For example, a 2019 article comparing several different COVID-19 molecular tests reported the following findings:

• The overall sensitivity of the tests was 95.9%, meaning there were very few false-negative results.
• The overall specificity was 97.2%, meaning there were very few false-positive results.
• Some tests were less accurate, with an overall sensitivity range of 80.5% to 96.6%.

A laboratory generally determines the accuracy of a test by measuring sensitivity and specificity. High levels of both measures mean that the test can correctly identify people who have a particular disease and those who do not.

Molecular genome testing can be challenging, and smaller mutations and genetic changes are generally more difficult to detect. The most popular methods detect only a small number of genetic problems.

Summary

Molecular testing is a type of genetic testing that provides important information about a person's genome. It can also detect signs of certain infections and genetic diseases based on the unique genetic characteristics that cause various diseases. This field is developing rapidly as more molecular testing techniques become available.

Genetic sequencing is advancing through next generation sequencing techniques. However, it is important for people to understand the limitations of molecular testing. This is particularly true for types of genetic testing with limited certainty, such as prenatal tests.