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Can You Trust Your Results? Verification of Spectrophotometers

Lab Academy

  • Photometry
  • Essay

Many laboratories are subject to regulatory requirements that e.g. are prescribed by Quality Management Systems. These laboratories are thus obligated to test work equipment and analysis instruments, including (spectro-) photometers, on a regular basis. What if such regulations do not apply to your lab? In this case, testing is not absolutely necessary, as it is not mandatory – but does that mean it is not important?

Anyone will tell you that, given a choice, they would prefer to avoid unnecessary effort, time and cost when it comes to laboratory experiments. Only a smooth, error-free performance of the entire experiment, from beginning to end, will guarantee accurate and reproducible results. This process includes analysis, as the quality of the data obtained for any sample is directly dependent on the quality of the measurement itself.

With respect to a standard laboratory application such as, for example, photometric quantification of nucleic acids, this principle would imply that, in addition to the purification procedure, the process of photometric measurement needs to be challenged and tested regularly. Errors may result from incorrect handling, or the cause may be of a technical nature. By employing a suitable troubleshooting approach with “step-by-step” error analysis, application errors are detected with relative ease [1, 2]. In contrast, many technological errors can only be uncovered with the help of comparative measurements and reference materials [3, 4].

How do technological errors even arise? The use of photometers over an extended period of time may lead to signs of wear that result from mechanical stress. In addition, different environmental factors (e.g. temperature, humidity, dust) may impact performance. Then, there is always the risk of damage or contamination through use or transport. Since photometry is a sensitive measurement technology, incorrect results may thus ensue. In order to reliably identify errors of such nature, it is crucial that photometers are tested on a regular basis.

Checking for visible damage and dirt, especially inside the cuvette shaft, constitutes the first step. If the instrument is equipped with an integrated self-test function, certain basic functions can also be tested at regular intervals or as needed (figure 1a).

In order to gain detailed information on the accuracy and precision of the measured values, photometric testing of the (spectro-)photometer is essential. To this end, measurements are carried out using certified, traceable reference materials (figure 1b). These commonly comprise filter sets with defined properties which are capable of establishing the wavelengths and the photometric accuracy of an instrument. The values obtained by applying the filters are then compared to the nominal values of the reference standards, and this comparison allows the user to determine whether the instrument performs within technical specifications. It should be noted that the reference materials themselves must be tested and certified on a regular basis.

In-depth inspection of a photometer can only be performed by a qualified service firm. Such service encompasses adjustments, repair and certification, if required.
Figure 1: Two methods for inspecting an Eppendorf BioSpectrometer®
a) Result of an instrument self-test of the Eppendorf BioSpectrometer kinetic
b) Result of the test of photometric accuracy of an Eppendorf BioSpectrometer using the associated reference filter set

Regular monitoring of photometers can ensure that their specifications continue to be met, even after long service. Appropriate verification methods will detect and eliminate sources of error, thus safeguarding the quality of analysis results.