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Common rhetoric over the past 30 or so years to do with performance-enhancing drugs is that the authorities are not keeping up with the cheaters. Not so true anymore.

Currently, the World Anti-Doping Agency (WADA) and the Athletics Integrity Unit have implemented out-of-competition testing in places throughout the world where it was previously difficult to do so. They also make good use of the Athlete Biological Passport, which monitors changes in athletes’ blood profiles over time. When dramatic changes occur, the alarm bells go off. There is also the whereabouts program, where certain athletes must be available for testing at pre-established windows of time. If they fail to be available three times in a twelve-month period, the incident is considered equal to a positive drug test and an appealable ban is handed down.

The next step in the efforts to stay in front of the athletes and their doping may be an advancement in blood sample storage technology, in particular a process called Dried Blood Spot (DBS) or Blood Spot Testing.

At the University of Victoria, a team of six scientists and lab technicians were considered to be part of a global revolution back in January 2016. At the time, it was a new, less-invasive method that could be applied to a variety of diseases and wellness monitoring.

DBS is a form of collection where patients place blood drops on a filter card after a finger prick using a lancet. Once dry, blood spot cards are stable for shipment and long-term storage.

How is it being used in anti-doping?

Dr. Marc Luginbühl, an analytical chemist who focuses on the analysis of direct alcohol biomarkers, illicit drug screening, and doping detection is a leading scientist in DBS. He recently published a scientific perspective article on DBS (as well as 15 others).  

His research currently targets this approach for this minimally invasive sampling procedure. He is currently working for the CAMAG DBS laboratory and serves as the president of The Society of Phosphatidylethanol Research (PEth-NET).

CAMAG is a Swiss-based company with a 60-year history as a world leader in instrumental High-Performance Thin-Layer Chromatography (HPTLC).

Athletics Illustrated interviewed Dr. Luginbühl about DBS. Here, he clarifies for readers the Dried Blood Spot procedure, which will enable anti-doping agencies to better store and test blood samples taken from athletes. 

The interview

Christopher Kelsall: Congratulations on getting the paper “Dried Blood Spots for Anti-Doping: Why Just Going Volumetric May Not Be Sufficient” accepted for publication in the Journal Drug Testing and Analysis.

From a non-scientific perspective, it sounds like dried blood spot testing is literally just that: dried blood, whereas the regular volumetric blood is in a beaker in liquid form, yes?

Marc Luginbühl: From a non-scientific perspective, this is true. From a scientific perspective, the term dried blood spots involve much more than just drying blood.

Nowadays, it has become a synonym for a process of minimally invasive blood sampling and applying it onto a supportive material. The supportive material can be either a filter paper (cellulose membrane) or a hydrophilic porous material and is commonly part of a sampling device. Once the sample has reached the analytical laboratory, the dried blood needs to be extracted to perform anti-doping analysis. Simply put, the sample preparation within the laboratory brings the DBS back into a liquid form, whereby we can selectively extract and analyze certain analytes.

CK: The process of sample collection and analysis appears to be closely connected. However, the WADA decided to split these processes and build two working groups.

ML: Absolutely. However, these areas are deeply connected, because the choice of sampling device determines the analytical process within the laboratory. The most convenient sampling device for the athlete can lead to significantly more work in the laboratory.

My major concerns here are about sample traceability and automation. For example, when using sponges manufactured from a hydrophilic porous material (as embedded within some push-button blood collection devices), the connection between the sample identification code and the sample itself is lost once the sampling device is opened and the samples are accessed for analysis. Also, getting those sponges out of the device and into a liquid extract requires many manual steps. When using DBS cards, the sample extraction can be performed without losing the connection between the sampling device and the sample itself. The cards can be shipped with standard mail and are a well-known medium for conserving dried blood spots for over 50 years. Furthermore, sample extraction and analysis of cards can be fully automated, which will be a key feature for the anti-doping DBS implementation.

Dr. Marc Luginbühl

CK: Why is DBS a better way to test for performance-enhancing drugs?

 ML: Dependent on the sampling device and the strategy (replacing or complementary to existing blood and urine sampling), the use of DBS brings advantages such as minimally invasive sampling, reduced sampling costs, easier shipment and storage, as well as automated laboratory processes. Also, the blood sampling frequency could be increased, as DBS sampling is more convenient for the athletes. I see the implementation of DBS into the world of anti-doping as a very important step forward.

You have approached the WADA with this article. How soon should they be able to use this process for testing samples?

ML: The laboratory processes to perform reliable hematocrit measurements from DBS, as suggested within the scientific article, are already available today. What I am still waiting for are reliable DBS guidelines which also account for the necessity of the hematocrit measurement when it comes to quantitative DBS sampling. Furthermore, for the DBS strategy to become a success story, a sampling device which can be used worldwide is necessary. This is mainly a regulatory question; as medical devices need to be registered. This is the case for standard DBS cards, as most countries use them for newborn screening. Additional sampling tools such as blood spot lancets and alcohol pads for disinfection can be sourced locally.

Currently, various anti-doping organizations are experimenting with novel DBS sampling strategies and devices. However, as soon as embedded needles or microneedles such as the ones within the push-button blood collection devices are involved, regulation will become a major hurdle. Although these devices are FDA-approved, it is questionable whether they will ever be globally approved.

CK: For in-competition tests, some athletes have already allowed the PEDs to flush out of the system so that they test clean and or micro-dose, which apparently also allows quicker flushing. How will DBS help in that case?

ML: The WADA has blood collection guidelines in place. These state that blood samples should not be collected within two hours of training or competition where athletic biological passport testing is to be conducted or within 30 minutes of training or competition where the analysis for growth hormone (GH) is to be conducted. The mentioned case will not change without a revision of these guidelines for DBS sampling.

CK: Will DBS extend the life of samples so that testing can be done later than the current eight years? I assume sample storage will be less expensive.

ML: The process of drying blood on a substrate immobilizes and thereby inactivates enzymes that are present within the blood. This inactivation stops the formation or degradation of certain analytes. In many studies, this has been reported as an advantage compared to conventional blood sampling. The overall stability of a DBS sample is dependent on the target analyte, the drying process, and the chosen storage condition. Usually, a reduction in storage temperature is beneficial for the overall sample stability. Due to their reduced size, DBS could be stored cost-efficiently at -80°C for years.

Read the study here>>