Molecular - Draeger

Molecular

The Dr?ger DrugTest 5000 can reveal if someone is under the influence of drugs in a matter of minutes. Its precision is based on SPECIFIC ANTIBODIES ? they fit the intoxicants like a key in a lock.

Text: Dr. Hildegard Kaulen

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EVIDENCE OF DRUGS

IMMUNE DIAGNOSTICS

D

Cannabis sativa is the Latin name for

hemp. The picture shows a shoot axis under

dark-field microscopy

Drug use is a social reality. Many people cannot and no longer want to go without the mind-altering kick, even though it may be at the cost of their health sooner or later. According to a recent report published by the Federal Government, one in four Germans aged between 18 and 64 has used cannabis; 319,000 German citizens are dependent on illegal drugs. The accident statistics from the Federal Statistics Office are also very clear: in 2013 no less than 3,896 road accidents involved people under the influence of drugs; 1,913 people were injured, 457 of them seriously. Several dozen people died because they or others had taken drugs shortly beforehand.

If law enforcement officers wish to test a person's fitness to drive, it has to be done quickly and the test must provide information about the current situation. "The police have to measure drug

use at that moment in time, not past use," says Rainer Polzius, responsible for the development of the DrugTest 5000 at Dr?ger. "It's not about establishing whether a driver was fit to drive yesterday or last week, but whether they are fit to drive now!" The device tests saliva and not blood or urine. Drugs do not last long in saliva, but can be detected in urine for a considerably longer period of time. Tetrahydrocannabinol (THC) COOH (carboxylic acid), the main metabolite of THC, can even be traced weeks later. Saliva, on the other hand, is easy to obtain during a roadside test. In addition, the sample cannot be manipulated ? any attempt at deception would be immediately noticed.

DR?GER REVIEW 111 | 2 / 2015

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PHOTO: HANS-J?RGEN KOCH

IMMUNE DIAGNOSTICS

EVIDENCE OF DRUGS

Antibodies are molecular heroes when testing for the presence of drugs

Dr?ger DrugTest 5000 ? the presence of eight commonly abused substances can currently be established using specific antibodies. These include cannabis and amphet-

amines, the most commonly used drugs, according to the World Drug Report 2014

The saliva test is based on an antigen?antibody reaction, similar to a pregnancy test. Antibodies are an integral part of immune diagnostics due to their high specificity. You could also call them molecular heroes. The immune system defends itself against all impurities with antibodies and does so very precisely, only forming the kind of antibodies which precisely match the foreign models. If antibodies encounter a specific antigen, an immune complex forms. The specific identification is also the reason for the hero status of the antibodies, because this is the basis for the broad value of this molecule class. In laboratory diagnostics, antibodies are used to detect the presence of proteins, hormones, toxins, or infectious agents. In medicine, chronically inflamed conditions and cancer are treated with antibodies, because they precisely target the area requiring therapy. Vaccines are also based on the efficacy of antibodies: in active immunization, an immunological memory is built up out of antibodies produced by the person's immune system; in the passive form doctors help in an acute situation by administering suitable antibodies directly into the blood. The heroic act is always the specific identification of the antigen.

The DrugTest 5000 uses drug-specific antibodies to prove the presence of more than half a dozen drugs: amphetamines, meth-

amphetamines, opiates, cocaine, benzodiazepines, cannabis, the heroin substitute methadone, and the synthetic narcotic ketamine. The only problem is trying to differentiate between the remnants of a poppy seed roll and heroin in the saliva, which is also extremely difficult chemically. Drug-specific antibodies react to a group of active agents, allowing several representatives of the same class of drugs to be identified at the same time. The saliva test consists of a sample collector (the actual test cassette with the testing strips), a buffer cartridge, and the analysis device itself. The test cassette contains five strips, each with two positions to ascertain the presence of drugs. "It is possible to prove the presence of up to ten intoxicants," says Rainer Polzius. "At the moment, eight positions are occupied, but we are continually examining the relevance and technical possibilities of proving the presence of new drugs."

Competition as a testing principle

The testing strips are made from a porous membrane; the test itself functions automatically. Once the saliva sample, test cassette, and buffer cartridge have been inserted into the analysis device, the internal temperature is brought to the optimal level and the saliva is mixed with the buffer solution contained within the buffer cartridge. The testing strips are then dipped into this mixture and the liquid migrates in one direction across the strips by capillary force. This is when what is also known in technical terms as the "lateral flow test" begins: the drug-specific antibodies are positioned on the membrane just after the starting point; located in the middle of each testing strip is a small amount of the drug whose presence is to be tested. When the mixture of saliva and buffer solution moves

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DR?GER REVIEW 111 | 2 / 2015

Methamphetamine forms crystals

resembling ice. The image shows

these under polarized light

microscopy

PHOTOS: DR?GERWERK AG & CO. KGAA, DENNIS KUNKEL MICROSCOPY, INC./VISUALS UNLIMITED/CORBIS

A little lexicon of molecules

Living beings protect themselves from foreign substances and pathogens using their immune systems. Its most important protagonists also help in the area of immune diagnostics.

Antigen = A substance identified by the immune system as foreign and responsible for the formation of antibodies.

Antibodies = Proteins formed by the B cells of the immune system. They identify antigens and help to fend them off as part of an immune response.

B cells = Immune cells which are part of white blood cells and form antibodies.

B cell clone = A B cell clone is a group of identical B cells produced by the division of one single B cell.

Epitope = The part of the antigen identified by the antibody and to which the antibody specifically attaches itself.

Immune diagnostics = The area of diagnostics which identifies substances or molecules using the high specificity of an antigen?antibody reaction. This is established by means of markers on the antibodies. Monoclonal antibodies are generally used.

Monoclonal antibodies are produced by a single clone of B cells and identify just one single epitope of the antigen. Monoclonal antibodies are identical among each other.

Polyclonal antibodies are produced by many different clones of B cells and identify various epitopes of the antigen. Polyclonal antibodies are a mixture of various antibodies.

Lock and key theory = The principle which states that two or more complementary structures can only fulfill their biological function if they fit together precisely. The identification of an antigen by an antibody follows this principle.

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IMMUNE DIAGNOSTICS

EVIDENCE OF DRUGS

Rapid test in less than ten minutes

across the testing strips, the drug-specific antibodies are released from the membrane and taken with it. If the saliva contains drugs, the molecules immediately combine with the relevant antibody. This is then no longer able to bond with the reference substance located in the middle, because it is already occupied with the drug molecules from the saliva sample. If there are no drugs present, the following happens: the antibodies attach themselves to the reference substances located in the middle, because they have not reacted with the drug molecules from the saliva. The test principle is thus a competing reaction. The visible proof is in the form of a marker on the drug-specific antibodies. This involves gold particles which emit light of a certain wavelength as soon as they themselves are exposed to light. If the analysis device registers a light signal, the antibody has attached itself to the drug in the middle of the testing strip. This means that no drug molecules are present in the saliva. If no light signal is registered, this is almost certainly proof of the presence of drugs. There is thus an inverse relationship between the drug concentration in the saliva and the signal strength via the antibody bonded in the middle of the testing strip. After five minutes, the results for seven drugs are available ? all except cannabis. Testing for the presence of tetrahydrocannabinol takes around three minutes longer. The DrugTest 5000 is a presumptive test device which predicts current drug use with a high degree of probability. Subsequent tests have to be performed in the laboratory for confirmation of the presumptive test. Laboratory analysis is necessary due to the potential for cross-reactivity or mouth contaminants.

Drug tests in the workplace

The saliva test can not only be used to assess a person's fitness to drive, but also to test drug use in the workplace or in jails. In Germany, drug tests in the workplace are a rarity, because they infringe on the personal rights of the employee. At the same time, there are many professions with a high accident risk where drug use among employees can have devastating consequences. These include com-

mercial drivers, pilots, forklift truck drivers, and crane operators, as well as the nuclear energy and (petro)chemical industries. "Drug tests in Germany are voluntary or clearly governed by an employment agreement," explains Rainer Polzius. "Following a judgment by an industrial tribunal in Hamburg, however, reasonable suspicion is not necessary if the activity is associated with considerable danger and the fitness for duty cannot be established in any other way." It is different in the United States, Australia, and other European countries. In these countries safety comes before the personal rights of employees. Ultimately, most people's drug abuse goes unnoticed, because they do not conform to the common clich? of a drug user.

Risk: new drugs

The drug market is always being flooded with new psychoactive substances. These are derivatives of approved medication, natural substances, or compositions which the pharmaceutical industry synthesized at one point and then later abandoned. None of the products have ever been rigorously tested. The side effects, toxicity, and dispersal in the body are only gradually learned from the emergency rooms or autopsy reports in the area of forensic medicine. Dealing and possessing the new drugs is a criminal offense if they are covered by the German Controlled Substances Act (BtMG). For some of them this is the case; for many of them it is not. This is either because their existence is not even known or because the process takes time. As soon as the

substances are listed in the BtMG, drug makers modify the basic chemical structure and thus circumvent the law. The whole thing is like the tale of the tortoise and the hare, in which the drug makers (mostly) have their noses in front. One or two new designer drugs appear every week.

PHOTO : STAR-MEDIA

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