Bioassay

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Bioassay setup
Microplastic toxicity assay in Daphnia magna - 1-s2.0-S0269749120360802-gr2 lrg.jpg
A biological test system (here: Daphnia magna ) is exposed to various experimental conditions (here: several microplastics preparations), to which it reacts.
MTT Plate.jpg
Some indicator of these reactions (e.g. a color change) is assessed, typically in a highly automated fashion through microplates like this.

A bioassay is an analytical method to determine the potency or effect of a substance by its effect on living animals or plants (in vivo), or on living cells or tissues (in vitro). [1] [2] A bioassay can be either quantal or quantitative, direct or indirect. [3] If the measured response is binary, the assay is quantal; if not, it is quantitative. [3]

Contents

A bioassay may be used to detect biological hazards or to give an assessment of the quality of a mixture. [4] A bioassay is often used to monitor water quality as well as wastewater discharges and its impact on the surroundings. [5] It is also used to assess the environmental impact and safety of new technologies and facilities.[ citation needed ]

Principle

A bioassay is a biochemical test to estimate the potency of a sample compound. Usually this potency can only be measured relative to a standard compound. [3] [1] A typical bioassay involves a stimulus (ex. drugs) applied to a subject (ex. animals, tissues, plants). The corresponding response (ex. death) of the subject is thereby triggered and measured. [6]

History

The first use of a bioassay dates back to the late 19th century, when the foundation of bioassays was laid down by German physician Paul Ehrlich. [7] He introduced the concept of standardization by the reactions of living matter. [7] [6] His bioassay on diphtheria antitoxin was the first bioassay to receive recognition. [8] His use of bioassay was able to discover that administration of gradually increasing dose of diphtheria in animals stimulated production of antiserum. [9]

One well known example of a bioassay is the "canary in the coal mine" experiment. [10] To provide advance warning of dangerous levels of methane in the air, miners would take methane-sensitive canaries into coal mines. If the canary died due to a build-up of methane, the miners would leave the area as quickly as possible.

Many early examples of bioassays used animals to test the carcinogenicity of chemicals. [11] In 1915, Yamaigiwa Katsusaburo and Koichi Ichikawa tested the carcinogenicity of coal tar using the inner surface of rabbit's ears. [11]

From the 1940s to the 1960s, animal bioassays were primarily used to test the toxicity and safety of drugs, food additives, and pesticides. [11]

Beginning in the late 1960s and 1970s, reliance on bioassays increased as public concern for occupational and environmental hazards increased. [11]

Classifications

Bioassay can be classified by how it is applied and how the response is recorded.

Direct assay
In a direct assay, the stimulus applied to the subject is specific and directly measurable, and the response to that stimulus is recorded. The variable of interest is the specific stimulus required to produce a response of interest (ex. death of the subject). [6] [12]
Indirect assay
In an indirect assay, the stimulus is fixed in advance and the response is measured in the subjects. The variable of interest in the experiment is the response to a fixed stimulus of interest. [6] [12]
Quantitative response
The measurement of the response to the stimulus is on a continuous scale (ex. blood sugar content, degree of color change in cell growth medium). [12]
Quantal response
The response is binary; it is a determination of whether or not an event occurs (ex. death of the subject). [12]

Examples

Ames test procedure Ames test.svg
Ames test procedure

One classical bioassay is the Ames test. A strain of Salmonella that requires histidine to grow is put on two plates with growth medium containing minimal amounts of histidine and some rat liver extract (to mimick liver metabolism). A suspected mutagen is added to one plate. If the plate with the suspected mutagen grows more visible colonies, it is probably mutagenic: a mutagen might cause the strain of bacterium to regain the ability to make its own histidine. [13]

Most other forms of toxicology testing are also bioassays. Animals or cell cultures may be put under a number of levels of a suspected toxin to ascertain whether the substance causes harmful changes and at what level it does so. The LD50 value, a common measure of acute toxicity, describes the dose at which a substance is lethal to 50% of tested animals. [14]

The potency of a drug may be measured using a bioassay. [15]

Environmental bioassays

Environmental bioassays are generally a broad-range survey of toxicity. [16] A toxicity identification evaluation is conducted to determine what the relevant toxicants are. Although bioassays are beneficial in determining the biological activity within an organism, they can often be time-consuming and laborious. Organism-specific factors may result in data that are not applicable to others in that species. For these reasons, other biological techniques are often employed, including radioimmunoassays. See bioindicator.

Water pollution control requirements in the United States require some industrial dischargers and municipal sewage treatment plants to conduct bioassays. These procedures, called whole effluent toxicity tests, include acute toxicity tests as well as chronic test methods. [5] The methods involve exposing living aquatic organisms to samples of wastewater for a specific length of time. [17] [18] Another example is the bioassay ECOTOX, which uses the microalgae Euglena gracilis to test the toxicity of water samples. [19] (See Bioindicator#Microalgae in water quality)

See also

Related Research Articles

<span class="mw-page-title-main">Ames test</span> Biological testing method

The Ames test is a widely employed method that uses bacteria to test whether a given chemical can cause mutations in the DNA of the test organism. More formally, it is a biological assay to assess the mutagenic potential of chemical compounds. A positive test indicates that the chemical is mutagenic and therefore may act as a carcinogen, because cancer is often linked to mutation. The test serves as a quick and convenient assay to estimate the carcinogenic potential of a compound because standard carcinogen assays on mice and rats are time-consuming and expensive. However, false-positives and false-negatives are known.

<span class="mw-page-title-main">Mutagen</span> Physical or chemical agent that increases the rate of genetic mutation

In genetics, a mutagen is a physical or chemical agent that permanently changes genetic material, usually DNA, in an organism and thus increases the frequency of mutations above the natural background level. As many mutations can cause cancer in animals, such mutagens can therefore be carcinogens, although not all necessarily are. All mutagens have characteristic mutational signatures with some chemicals becoming mutagenic through cellular processes.

<span class="mw-page-title-main">Daminozide</span> Chemical compound

Daminozide, also known as aminozide, Alar, Kylar, SADH, B-995, B-nine, and DMASA, is a plant growth regulator. It was produced in the U.S. by the Uniroyal Chemical Company, Inc,, which registered daminozide for use on fruits intended for human consumption in 1963. In addition to apples and ornamental plants, they also registered it for use on cherries, peaches, pears, Concord grapes, tomato transplants, and peanut vines. Alar was first approved for use in the U.S. in 1963. It was primarily used on apples until 1989, when the manufacturer voluntarily withdrew it after the U.S. Environmental Protection Agency proposed banning it based on concerns about cancer risks to consumers.

<span class="mw-page-title-main">Toxicity</span> Degree of harmfulness of substances

Toxicity is the degree to which a chemical substance or a particular mixture of substances can damage an organism. Toxicity can refer to the effect on a whole organism, such as an animal, bacterium, or plant, as well as the effect on a substructure of the organism, such as a cell (cytotoxicity) or an organ such as the liver (hepatotoxicity). By extension, the word may be metaphorically used to describe toxic effects on larger and more complex groups, such as the family unit or society at large. Sometimes the word is more or less synonymous with poisoning in everyday usage.

In vitro toxicity testing is the scientific analysis of the toxic effects of chemical substances on cultured bacteria or mammalian cells. In vitro testing methods are employed primarily to identify potentially hazardous chemicals and/or to confirm the lack of certain toxic properties in the early stages of the development of potentially useful new substances such as therapeutic drugs, agricultural chemicals and food additives.

<span class="mw-page-title-main">Piperonyl butoxide</span> Chemical compound

Piperonyl butoxide (PBO) is a pale yellow to light brown liquid organic compound used as a synergist component of pesticide formulations. That is, despite having no pesticidal activity of its own, it enhances the potency of certain pesticides such as carbamates, pyrethrins, pyrethroids, and rotenone. It is a semisynthetic derivative of safrole.

An assay is an investigative (analytic) procedure in laboratory medicine, mining, pharmacology, environmental biology and molecular biology for qualitatively assessing or quantitatively measuring the presence, amount, or functional activity of a target entity. The measured entity is often called the analyte, the measurand, or the target of the assay. The analyte can be a drug, biochemical substance, chemical element or compound, or cell in an organism or organic sample. An assay usually aims to measure an analyte's intensive property and express it in the relevant measurement unit.

Genotoxicity is the property of chemical agents that damage the genetic information within a cell causing mutations, which may lead to cancer. While genotoxicity is often confused with mutagenicity, all mutagens are genotoxic, but some genotoxic substances are not mutagenic. The alteration can have direct or indirect effects on the DNA: the induction of mutations, mistimed event activation, and direct DNA damage leading to mutations. The permanent, heritable changes can affect either somatic cells of the organism or germ cells to be passed on to future generations. Cells prevent expression of the genotoxic mutation by either DNA repair or apoptosis; however, the damage may not always be fixed leading to mutagenesis.

<span class="mw-page-title-main">Auxotrophy</span> Inability to synthesize an organic compound required for growth

Auxotrophy is the inability of an organism to synthesize a particular organic compound required for its growth. An auxotroph is an organism that displays this characteristic; auxotrophic is the corresponding adjective. Auxotrophy is the opposite of prototrophy, which is characterized by the ability to synthesize all the compounds needed for growth.

<span class="mw-page-title-main">Aquatic toxicology</span> Study of manufactured products on aquatic organisms

Aquatic toxicology is the study of the effects of manufactured chemicals and other anthropogenic and natural materials and activities on aquatic organisms at various levels of organization, from subcellular through individual organisms to communities and ecosystems. Aquatic toxicology is a multidisciplinary field which integrates toxicology, aquatic ecology and aquatic chemistry.

<span class="mw-page-title-main">Bioindicator</span> Species that reveals the status of an environment

A bioindicator is any species or group of species whose function, population, or status can reveal the qualitative status of the environment. The most common indicator species are animals. For example, copepods and other small water crustaceans that are present in many water bodies can be monitored for changes that may indicate a problem within their ecosystem. Bioindicators can tell us about the cumulative effects of different pollutants in the ecosystem and about how long a problem may have been present, which physical and chemical testing cannot.

<span class="mw-page-title-main">Ecotoxicology</span>

Ecotoxicology is the study of the effects of toxic chemicals on biological organisms, especially at the population, community, ecosystem, and biosphere levels. Ecotoxicology is a multidisciplinary field, which integrates toxicology and ecology.

<span class="mw-page-title-main">Wastewater quality indicators</span> Ways to test the suitability of wastewater

Wastewater quality indicators are laboratory test methodologies to assess suitability of wastewater for disposal, treatment or reuse. The main parameters in sewage that are measured to assess the sewage strength or quality as well as treatment options include: solids, indicators of organic matter, nitrogen, phosphorus, indicators of fecal contamination. Tests selected vary with the intended use or discharge location. Tests can measure physical, chemical, and biological characteristics of the wastewater. Physical characteristics include temperature and solids. Chemical characteristics include pH value, dissolved oxygen concentrations, biochemical oxygen demand (BOD) and chemical oxygen demand (COD), nitrogen, phosphorus, chlorine. Biological characteristics are determined with bioassays and aquatic toxicology tests.

<span class="mw-page-title-main">Dose–response relationship</span> Measure of organism response to stimulus

The dose–response relationship, or exposure–response relationship, describes the magnitude of the response of an organism, as a function of exposure to a stimulus or stressor after a certain exposure time. Dose–response relationships can be described by dose–response curves. This is explained further in the following sections. A stimulus response function or stimulus response curve is defined more broadly as the response from any type of stimulus, not limited to chemicals.

<span class="mw-page-title-main">Triclocarban</span> Antimicrobial agent

Triclocarban is an antibacterial chemical once common in, but now phased out of, personal care products like soaps and lotions. It was originally developed for the medical field. Although the mode of action is unknown, TCC can be effective in fighting infections by targeting the growth of bacteria such as Staphylococcus aureus. Additional research seeks to understand its potential for causing antibacterial resistance and its effects on organismal and environmental health.

<span class="mw-page-title-main">SOS chromotest</span>

The SOS chromotest is a biological assay to assess the genotoxic potential of chemical compounds. The test is a colorimetric assay which measures the expression of genes induced by genotoxic agents in Escherichia coli, by means of a fusion with the structural gene for β-galactosidase. The test is performed over a few hours in columns of a 96-well microplate with increasing concentrations of test samples. This test was developed as a practical complement or alternative to the traditional Ames test assay for genotoxicity, which involves growing bacteria on agar plates and comparing natural mutation rates to mutation rates of bacteria exposed to potentially mutagenic compounds or samples. The SOS chromotest is comparable in accuracy and sensitivity to established methods such as the Ames test and is a useful tool to screen genotoxic compounds, which could prove carcinogenic in humans, in order to single out chemicals for further in-depth analysis.

For pharmacology and genetics, the Umu Chromotest, first developed and published by Oda et al., is a biological assay (bioassay) to assess the genotoxic potential of chemical compounds. It is based on the ability of DNA-damaging agents to induce the expression of the umu operon. In connection with the damage inducible (din) genes recA, lexA and umuD, the umuC gene is essentially involved in bacterial mutagenesis through the SOS response.

An early life stage (ELS) test is a chronic toxicity test using sensitive early life stages like embryos or larvae to predict the effects of toxicants on organisms. ELS tests were developed to be quicker and more cost-efficient than full life-cycle tests, taking on average 1–5 months to complete compared to 6–12 months for a life-cycle test. They are commonly used in aquatic toxicology, particularly with fish. Growth and survival are the typically measured endpoints, for which a Maximum Acceptable Toxicant Concentration (MATC) can be estimated. ELS tests allow for the testing of fish species that otherwise could not be studied due to length of life, spawning requirements, or size. ELS tests are used as part of environmental risk assessments by regulatory agencies including the U.S. Environmental Protection Agency (EPA) and Environment Canada, as well as the Organisation for Economic Co-operation and Development (OECD).

Toxicological databases are large compilations of data derived from aquatic and environmental toxicity studies. Data is aggregated from a large number of individual studies in which toxic effects upon aquatic and terrestrial organisms have been determined for different chemicals. These databases are then used by toxicologists, chemists, regulatory agencies and scientists to investigate and predict the likelihood that an organic or inorganic chemical will cause an adverse effect on exposed organisms.

<span class="mw-page-title-main">CompTox Chemicals Dashboard</span> Chemical database

The CompTox Chemicals Dashboard is a freely accessible online database created and maintained by the U.S. Environmental Protection Agency (EPA). The database provides access to multiple types of data including physicochemical properties, environmental fate and transport, exposure, usage, in vivo toxicity, and in vitro bioassay. EPA and other scientists use the data and models contained within the dashboard to help identify chemicals that require further testing and reduce the use of animals in chemical testing. The Dashboard is also used to provide public access to information from EPA Action Plans, e.g. around perfluorinated alkylated substances.

References

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