A common way to assess the effectiveness of cleaning and sanitation programs in food manufacturing facilities is through the use of methods that detect adenosine triphosphate (ATP). Methods based on ATP detection are inexpensive and rapid, and provide the ability to perform onsite in real-time. There are several manufacturers of ATP-based methods, but choosing the most reliable one can be a daunting task. This article will discuss how these methods work and which factors should be considered to make an informed purchasing decision.

ATP is the universal energy currency in all living cells. It is present in all viable microorganisms (with the exception of viruses) and in foodstuffs. High amounts of ATP can be found in some fresh foods like vegetables, while other foods, especially highly processed foods such as fats, oils or sugar, contain very low amounts of this molecule. It is also important to know that ATP can be found in the environment in its free form hours after a cell has died.¹ An ATP bioluminescence assay operates on the principle that ATP in food/food residues and microorganisms, in the presence of a luciferin/luciferase complex, leads to light emission. This light can be measured quantitatively by a luminometer (light-detecting instrument), with results available in 10–40 seconds. The amount of light emitted is proportional to the amount of ATP on a surface and hence its cleanliness. The light emitted is typically measured in relative light units (RLUs), calibrated for each make of instrument and set of reagents. Therefore, the readings obtained from assessing the cleaning of food manufacturing facilities need to be compared with baseline data representing acceptable clean values.

Varying Optical Components

Luminometers have evolved over the years from very large and cumbersome in size to small handheld models that can be used anywhere within a manufacturing facility. Although several components are housed inside these instruments, the optical component is the most important part of a luminometer. Used to detect light coming from the ATP/luciferin/luciferase reaction, the optical component is the defining factor related to luminometer reliability, sensitivity and repeatability. Good luminometers use a photomultiplier tube (PMT) in the light detection system; however, as part of the drive toward cheaper and smaller instruments, some manufacturers have replaced PMTs with less-sensitive photodiode-based systems. When using photodiodes, the swab chemistry must be adapted to produce more intense light. This results in a shorter duration of light, decreasing the time window allotted to place the swab in the luminometer and obtain an accurate read. A PMT, however, multiplies the electrical current produced when light strikes it by millions of times, allowing this optical device to detect a single photon. This approach emits light over a longer period of time. Although the weight of the system is also dependent on factors such as the battery, case and the display screen, a luminometer constructed with a photodiode will generally weigh less than a luminometer constructed with a PMT, since the former is smaller than the latter.

Sensitivity Testing

When an ATP hygiene monitoring system has poor sensitivity or repeatability, there is substantial risk that the test result does not truly represent the hygienic status of the location tested. Therefore, it may provide false positives leading to unnecessary chemical and labor costs and production delays, or false negatives leading to the use of contaminated pieces of equipment. A system that is sensitive to low-level contamination of a surface by microorganisms and/or food residues allows sanitarians to more accurately understand the status of a test point. The ability of a system to repeat results gives one peace of mind that the result is reliable and the actions taken are appropriate. To test different ATP systems for sensitivity, one can run the following simple test using at least eight swabs per system:

•    Make at least four serial dilutions of a microbial culture and a food product in a sterile phosphate buffer solution.
•    Using an accurate pipette, dispense 20 μl of these dilutions carefully onto the tip of the swabs of each ATP system and read the swabs in the respective luminometer, following the manufacturer’s instructions.
•    Use caution when dispensing the inoculum onto the swab head to prevent any sample loss or spillage. In addition, it is very important the swabs are inoculated immediately prior to reading, which means that each swab should be inoculated one at a time and read in the respective luminometer. Repeat this process for all the swabs.

To test different ATP systems for sensitivity, one can run a simple test using at least eight swabs per system. Photo courtesy of 3M

The most sensitive system will be the one that results in the most “fail results” (using the manufacturers’ recommended pass/caution/fail limits).

One can also test different ATP systems for repeatability by the following test:

•    Prepare a dilution of a standard ATP positive control or a food product such as fluid milk in a sterile phosphate buffer. If using a standard ATP positive control, follow the manufacturer’s direction to prepare dilution. If using fluid milk, add 1 ml of milk into 99 ml of phosphate buffer.
•    Using an accurate pipette, dispense 20 μl of this standard onto the tip of the swabs of each ATP system and read these swabs in their respective luminometer, following the manufacturer’s instructions.
•    Prepare and read at least 10 swabs for each system you are evaluating, and capture the results on a digital spreadsheet.
•    Once all 10 swab results (for each system) are in the spreadsheet, calculate the mean (=average) and standard deviation (=stdev) for each system’s data set. Divide the standard deviation by the mean and transform the result in percentage; this value is called the coefficient of variation percentage (CV%).
The test with the lowest CV% is the most repeatable and will provide the most reliable information to help make the correct decisions for a food manufacturing facility.

Choosing the Right ATP System

There are many ATP systems available on the market to support cleaning and sanitation verification in manufacturing plants. Some systems are more reliable than others and will provide results that are meaningful, accurate and repeatable. Be sure, therefore, not to choose a system solely based on its price. Check for the quality of the instrument, ask the sales representative what kind of optical device is used in the construction of the instrument and, moreover, perform an evaluation running tests for both sensitivity and repeatability. It is also important to consider the functionality and usability of the software provided with the system to ensure that the software can be used to customize sample plans, store test results and produce reports and graphs.

Reference

1. Jay, J. M., ‎Loessner, M. J., & Golden, D. A. (2008). Modern Food Microbiology.

About the Author:

Camila Gadotti, M.S., is a field technical service professional and Michael Hughes is a technical service professional with 3M Food Safety.