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EIA Method for Detecting Drugs of Abuse and Therapeutic Drugs

Toxicology screening for drugs of abuse and therapeutic drugs is increasingly used in sectors beyond pharmaceutical, healthcare, and research industries, with a larger number of transportation, construction firms, employers, military and law enforcement agencies implementing drug testing policies to improve workplace safety.

Screening tests are performed by immunochromatographic or lateral flow assay, or by an enzyme immunoassay (EIA or ELISA), a biochemical method for detecting the presence of drugs of abuse in a biological sample. This is achieved through a combination of analyte specific antibody binding and colorimetric detection system.

We have described the immunochromatographic or lateral flow assay in a previous blog. We now will describe the principle of EIA for small analytes, such as therapeutic drugs, drugs of abuse, pesticides, certain toxins etc.

There are several EIA systems: Here we will use the detection of THC in urine as an example.

·         Competitive EIAs: In this assay, the analyte to be detected in the sample competes against an enzyme-labeled analyte for the available antibodies previously coated on the wells of the EIA plate. In the case of THC, the wells are coated with anti-THC antibody.  The sample containing THC is mixed with the enzyme-labeled THC (THC-HRP) then the mix is added to the wells and incubated. After equilibration of the reaction, the wells are rinsed to remove the unbound analyte, and the material that remains bound to the surface of the well is then treated with a substrate for the developing of color. The resultant chromatic properties of the solution allow for accurate quantitative analysis. The developing of color is directly proportional to the concentration of THC. The higher the concentration of THC present in the sample, the weaker the intensity of the color because of the competitive nature of the assay.

·         Sandwich assays. This method is not well suited for drug detections because it is difficult to have a sandwich assay with small molecules due to steric hindrance. For the THC detection example, wells are treated with an anti-THC antibody, then the sample to be tested is added to the well. After incubation, the wells are washed and a second anti-THC antibody conjugated to an enzyme is added to the sample. The THC in this example is already “swallowed” by the first antibody, so the second antibody will not be able to see it and the test will fail.

·         Direct EIA mechanisms use similar principles. In this EIA version, THC-BSA will be coated on the plate, the sample to be tested will be introduced next, followed by an anti-THC antibody conjugated to an enzyme.  The plate is incubated and subsequently a specific substrate is added to elicit a chromatic response.  In this case, stronger color changes are associated with higher presence of an antigen or specific drug in a sample.

Once a drug has metabolized in a biological system, its metabolites can be detected in varying concentrations in a range of samples from that system – the most common analytes for commercial wet chemistry toxicology screening are urine and oral fluids.

The complex and dynamic chemical characteristics of drugs of abuse presents significant challenges for accurate toxicology screening and drug detection in all sectors, with the increasing prevalence of synthetic variants of many drugs of abuse. Improved detection enzymes and antibodies with increased target specificity are required to accurately set biomarkers and laboratory cutoffs for new and emerging drugs.

Reagents for Detecting Drugs of Abuse and Therapeutic Drugs from Pyxis

Pyxis Laboratories designs and synthesizes standard and custom reagents for use in an increasing array of applications. Some of the conjugates and antigen-antibody paired reagents we provide for use in EIA or ELISA and lateral flow tests include:

·         Alcohol such as, ethyl glucuronide-BSA (EtG-BSA);

·         Amphetamines such as, Amphetamine-BSA, Methamphetamine-BSA, methcathinone-BSA, MDMA-BSA, MDPV-BSA, Mephedrone-BSA and Methylphenidate-BSA (Ritalin-BSA);

·         Benzodiazepines, such as Oxazepam-BSA and Clonazepam-BSA;

·         Cannabinoids such us, delta-8-THC-BSA, delta-9-THC-BSA; and synthetic cannabinoids such as, JWH-018-BSA, ABpinaca-BSA, UR144-BSA;

·         Cocaine, such as BZEG-BSA;

·         Hallucinogens/Anesthetics such as, Ketamine-BSA and PCP-BSA;

·         Nicotinic such as, Cotinine-BSA.

·         Opioids such as, Buprenorphine-BSA, Fentanyl-BSA, Methadone-BSA, Morphine-BSA, Oxycodone-BSA, Propoxyphene-BSA and others such as Chloramphenicol-BSA, Gabapentin-BSA and N-acetyl glucosamine-BSA (NAG-BSA).

These and many more conjugations allow for lot-to-lot reproducibility for consistent and accurate results applicable to an increasing range of applications. For any more information Pyxis products suitable for the detection of drugs of abuse, please contact us.