RADIO IMMUNOASSAY DANIEL CASAL

What is radioimmunoassay Sensitive method that measures small amounts of substances in the blood Able to measure drugs, antigens and hormones Any biological substance where an antibody exists can be measured Uses include drug detection, blood bank screening for certain viruses and cancers (such as HIV and Hepatitis B) Immunoassays produces signals in response to binding Able to be detected by many ways such as it showing a color change, fluoresces under light, or emitting radiation.

Radioimmunoassay relevance Most common method to test for drugs of abuse Screens presence of drugs in different areas such as forensic toxicology and clinical toxicology (Dated back from 1970 when soldiers were coming back from the Korean war) Take a urine or blood sample since its easy to collect and obtain in large quantity Radioimmunoassay technique isolated Diphenyldantoin from semen, blood stains saliva and dried saliva stains.

Radioimmunoassay Relevance Testing Opiates for morphine content is mostly performed in forensics urine drug test laboratories Doses of hydromorphine, hydrocodone, oxymorphone and oxycodone were administered to individuals. Urine sample were collected before and after the drugs were administered. The radioimmunoassay testing indicated that 3 of the 4 drugs were detectable by urine (Low to moderate concentrations of these drugs will go undetected when testing)

Method First obtain a substance that contains the antigen (containing antiserum) Introduce antigen with radioactive chemicals and then incubate it at +4C (Isotopes used are H3, I 125, I 131, C14 because they emit gamma rays). Most common is I 125 or I 131 (easier for Iodine atoms to be introduced in tyrosine residue) Mix the antibodies and the radioactive antigen Both substances bind with each other and end up forming a new substance Add unknown substance (Also known as “cold”) This causes the antigens in the new/unknown substance to bind with the antibodies

Method The unknown substance displace the isotope substance that was originally joined together with the antigen. If there is in increase in the cold antigen, there will be an increase in the radioactive antigen being displaced from the antibody molecule Able to measure radioactivity and a standard biding curve is shown Separate the bound from free antigen by precipitating the antigen antibody

Chemistry in radioimmunoassay Very useful in quantitative analysis of drugs. Antibodies reacting with specific elements to get a measurement Detection limit depends if the solution was in a liquid phase, or in a solid phase

Safety Always wear goggles, apron, and gloves at all times Be careful when handling the radioactive isotopes Stay away from the gamma rays being emitted during the radioimmunoassay process Dispose of the hazardous material accordingly

Problems/difficulties If your test tube isn’t cleaned then the sample will be contaminated Plug in wrong data Properties of specimens difficult to detect Interference Malfunction on the instrument used to count the radioactivity Radioactive isotope Antibodies and antigens aren’t pure

Limitations Really sensitive (smallest concentration of a substance that can be measured reliably Specificity depends on the antigen and the antibody and the isotope Contaminants affect both the precision and the accuracy of quantitative data Detection limits depend on the standard curves of the sample and it depends of the substance

Advantages Really sensitive and precise Able to detect small trace amounts Require small concentration of antigens Large capacity Disadvantages Usage of radioactive chemicals are extremely dangerous Only special trained individuals are allowed to perform the lab Require lots of arrangements and time to store/dispose radioactive material Expensive equipment Require pure antibodies and antigens Takes a lot of time

Bibliography http://www.txdps.state.tx.us/CrimeLaboratory/ http://www.randoxtoxicology.com/immunoassay-drug-testing http://www.ncbi.nlm.nih.gov/pubmed/7536861 http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/R/Radioimmunoassay.html http://www.slideshare.net/chakravarthyrapolu/radio-immuno-assay http://www.pharmatutor.org/articles/radiommunoassay-drugs-hormones?page=0,3 http://www.antibodies-online.com/resources/17/1215/Radioimmunoassay+RIA/ http://www.slideshare.net/banuman35/analytical-method-development-by- pravisankar http://europepmc.org/abstract/MED/709872