FOOD ANALYSIS: RAPID TESTING
Diagram
Two rapid detection tests
ELISA
Antibodies fixed to a plate catch the target microbe or its toxin. A second antibody linked to an enzyme makes a colour change that shows the target is present.
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PCR
The polymerase chain reaction copies a piece of the germ’s DNA until there is enough to detect, and names the germ in hours instead of days.
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The choice
ELISA finds a protein or toxin by antibody binding. PCR finds DNA, and says which genes the germ carries.
Figure 1: ELISA detects a target by antibodies and a colour change; PCR detects it by copying its DNA. Drawn in our own style from the FTN Food Microbiology notes, Unit 3, and the FTN Food Safety and Security notes, Unit 7.
Introduction
The enzyme-linked immunosorbent assay, written ELISA, and the polymerase chain reaction, written PCR, are the two rapid detection methods of food testing, and both answer in hours where a plate count takes days. The difference is the target: ELISA detects a protein, a microbe or its toxin, with antibodies, while PCR detects the DNA of the germ by copying it until there is enough to read. One finds the molecule, the other finds the gene.
The FTN Food Microbiology notes, Unit 3, describe ELISA for toxins and for screening, and the FTN Food Safety and Security notes, Unit 7, set PCR beside immunological tests, plate counts and ATP tests. This article follows those two units, with Pometto and colleagues, Food Biotechnology, chapter on ELISA assays for toxins in foods, and Sun, Handbook of Food Safety Engineering, Part II.
Quick comparison
Table 1: ELISA compared with PCR
| Aspect | ELISA | PCR |
|---|---|---|
| What it detects | A protein, a microbe or its toxin, caught by antibodies | A piece of the germ’s DNA |
| Target | The antigen or toxin itself | A DNA sequence, which can name the germ and its genes |
| How it works | Antibodies fixed to a plate or well catch the target from the sample, a second antibody linked to an enzyme binds to it, and the enzyme makes a colour change | The DNA is copied again and again until there is enough to detect |
| Answer given | Is this species or toxin present, and the strength of the colour can estimate how much is there | Is its DNA here, and which genes does it carry |
| Time | Hours, and suited to screening large numbers of samples | Hours instead of days, against days for a plate count |
| Before the test | Often needs an enrichment step: the sample is grown in broth first, so a rare target multiplies to a detectable level | Needs a clean sample, because food parts can block the reaction |
| Skill needed | A laboratory test that can be run on many samples at once | Needs a skilled worker |
| Best for | Toxins such as staphylococcal enterotoxin, and screening large numbers of samples for Salmonella or Listeria | Naming a germ such as Salmonella in a sample in hours, and finding which genes it carries |
| Limit | Salmonella may be present in very small numbers, below what the antibodies can detect, and food parts can block the reaction | It finds DNA, not living cells: a positive result may need a plate test to confirm, and the sample must be clean |
| Confirmation | A positive result may need a plate test to confirm it | A positive result may need a plate test to confirm it |
Scientific explanation
Definition
ELISA, the enzyme-linked immunosorbent assay, is a test in which antibodies fixed to a plate or well catch the target microbe or its toxin from the sample, a second antibody linked to an enzyme binds to it, and the enzyme makes a colour change: the colour shows the target is present, and its strength can estimate how much is there. PCR, the polymerase chain reaction, is a test that copies a piece of the germ’s DNA until there is enough to detect, and it can name the germ in hours instead of days.
Working principle
In ELISA, the first antibody is fixed to the plate and catches the target, the second antibody carries the enzyme to it, and the enzyme makes the colour change. The method is used for toxins such as staphylococcal enterotoxin and for screening large numbers of samples for Salmonella or Listeria. Because Salmonella may be present in very small numbers, below what the antibodies can detect, and food parts can block the reaction, ELISA often needs an enrichment step before testing: growing the sample in broth first multiplies the cells to a detectable level and dilutes the food parts that interfere. In PCR, a piece of the germ’s DNA is copied again and again until there is enough to detect, and the copy can name the germ and show which genes it carries. PCR is fast and exact but needs a clean sample and a skilled worker, while plate counts are slow but show living germs and their numbers, and rapid tests are fast but may need a plate test to confirm a positive result.
Apparatus
ELISA needs a plate or wells with antibodies fixed to them, a second antibody linked to an enzyme, and a reader for the colour change. PCR needs the reagents that copy the DNA, including primers that match the target sequence, and an instrument that runs the heating and cooling cycles. The books used here name no single apparatus list for either method beyond this, so no longer list is given.
Measurement procedure
Choosing a method means matching the question to the tool. A total count answers how many microbes are growing, a selective count answers how many of this group, ELISA or a strip answers is this species or toxin present, and PCR answers is its DNA here, and which genes does it carry. A plant that must release a batch today uses a fast method and confirms a positive with a culture method, because no single test is fast, cheap, specific and quantitative all at once. The Compendium of Methods for the Microbiological Examination of Foods gives standard procedures for each of these methods, so results from different labs can be compared.
Factors affecting readings
For ELISA, the number of target cells and the food matrix decide the result: very small numbers fall below what the antibodies can detect, and food parts can block the reaction, which is why enrichment comes first. For PCR, the cleanliness of the sample decides the result: food parts can block the copying, and the worker must be skilled. For both, a positive result may need a plate test to confirm it, because a rapid test is fast but a plate count shows living germs and their numbers.
Formulas and calculations
PCR copies double at each cycle, so after n cycles one starting copy has grown to 2 to the power n copies, in an ideal reaction. After 10 cycles that is about 1,000 copies, after 20 cycles about 1,000,000 copies, and after 30 cycles about 1,000,000,000 copies, which is why PCR can name a germ in hours instead of days.
For ELISA the books give no formula, so none is given: the strength of the colour change estimates how much target is present, read against standards.
Industrial applications
- ELISA: testing for toxins such as staphylococcal enterotoxin in foods.
- ELISA: screening large numbers of samples for Salmonella or Listeria.
- ELISA: answering is this species or toxin present, where a fast answer on many samples is needed.
- PCR: naming Salmonella in a sample in hours instead of days.
- PCR: finding which genes a germ carries, where the question is about its DNA.
- PCR: fast testing where a batch must be released today, with a plate test to confirm a positive result.
Advantages and limitations
- ELISA, advantage: it screens large numbers of samples at once, and the colour change shows the target and estimates how much is there.
- ELISA, limitation: it needs an enrichment step for rare targets, and food parts can block the reaction.
- PCR, advantage: fast and exact, naming the germ and its genes in hours instead of days.
- PCR, limitation: it needs a clean sample and a skilled worker, and it finds DNA, not living cells, so a positive may need a plate test to confirm it.
FAQ
What is the main difference between ELISA and PCR?
ELISA detects a protein, a microbe or its toxin with antibodies and a colour change, while PCR detects the DNA of the germ by copying it until there is enough to read.
Why does ELISA need an enrichment step before testing for Salmonella?
Salmonella may be present in very small numbers, below what the antibodies can detect, and food parts can block the reaction, so growing the sample in broth first multiplies the cells to a detectable level.
What does PCR tell the analyst that ELISA does not?
Which genes the germ carries, because PCR reads its DNA, while ELISA reads its proteins or toxins.
Why does a rapid test still need a plate test to confirm a positive?
Because no single test is fast, cheap, specific and quantitative all at once, and a plate count shows living germs and their numbers.
How fast is PCR compared with a plate count?
PCR names the germ in hours instead of days, because each cycle doubles the DNA, so a few copies grow to about a billion in 30 cycles.
References and further reading
- FTN subject: Food Microbiology, Unit 3: Analytical Techniques in Food Microbiology; FTN subject: Food Safety and Security, Unit 7: Microbiological Criteria, Sampling Plans and Testing.
- Anthony Pometto, Kalidas Shetty, Gopinadhan Paliyath and Robert E. Levin (eds.), Food Biotechnology, 2nd ed., CRC Press, 2005 (Google Books page read)
- Da-Wen Sun (ed.), Handbook of Food Safety Engineering, Wiley, 2011 (Google Books page read)
