Dear Dr. Susan Student Forum Discussion University of Cambridge First of all I am surprised that ELIZA has found its application in forensic science. ELISA is very familiar to us in diagnostic medicine and in medical research. It was first used in the 1970’s by medical scientists as a diagnostic tool for the detection of infectious diseases such as dengue, malaria, the detection of Mycobacterium antibodies in tuberculosis, in the diagnosis of hepatitis, the detection of HIV antibodies, and in serological blood test for coeliac disease as well as for the presence of enterotoxins produced by E. coli. It is also used in the study of immunology, in neuroscience, and in cancer research among many other biomedical applications. ELISA can also be used in toxicology as a rapid presumptive screen for certain classes of drugs, in allergic reactions in patients, or in serological testing for antigen-antibody reaction. Later ELIZA found its application in biotechnology, and in quality control of vaccine production and other enzyme-linked biological materials. ELIZA is also used in the quality control in the food industry in detecting potential food allergens, such as milk, peanuts, walnuts, almonds, and eggs during food manufacturing. All these developments are familiar to us All of them involve specific antigen- antibody reactions, and it is specific and sensitive only to certain type of agent we are looking for, although sometimes ELISA may give false positive and false negative results. Various types of toxins act differently because they are chemically different. For instance, a Spanish fly produces a toxin called cantharidin that can cause a type of blister. The hooded pitohui harbours a neurotoxin in its skin and feathers called homobatrachotoxin. The poison of a dart frog is different from that of a cane toad that produces bufotoxin So are snakes venoms which fall mainly into two broad classes of toxins. They are either, neurotoxins (mostly found in elapids) or haemotoxins (mostly found in viperids). Their toxicodynamics (mode of action of the toxins) are entirely different. Snake venoms act either on the nervous system or on the blood Toxins biological targets generally act either as binding proteins, or via ion channels, DNA, or a variety of other receptors. Because of their varying chemical structures and toxicodynamics, I don’t think ELISA can be used to detect all types of toxins and poisons that are not enzyme-linked. Some may not produce an antigen-antibody response In forensic toxicology I think if we suspect death or injury due to a biological venom, the first thing to do is to find out what bit the victim. Look for the lesion. Examine first its bite marks, history, features, morphology and presenting signs and symptoms (if victim is not dead) If we suspect a certain and specific poisonous animal, conduct only that specific test (ELISA, whatever) for that specific venom, else we would be searching for a needle in a haystack. On the second issue raised by another student, I do not think the degradation of any sample other than that specific ligand can interfere or give false positive or false negative to the results in an ELSA assay, unless we heat treat the ligand (protein) to destroy it. After all, we still have a positive and negative control to be doubly sure Furthermore, ELIZA is very simple to use, and is a very rapid test as most of the ELIZA test kits are already commercially-prepared and ready for use without the need for any elaborate laboratory. It can be done in the field. This is a step in advance in analytical chemistry over other wet chemical analysis (which conventionally of course can also be used, but is time consuming, and requires larger quantities of samples, besides specific analytical reagents). In normal wet chemistry, we need to reuse again and again the same apparatus after washing, whereas in ELISA the rapid kits can only be used once. Another analytical chemistry approach in toxicology, is to use various types of sophisticated spectrophotometric analysis. There are many types of spectrophotometers for various chemical analysis. They range anything from absorption spectroscopy, ultraviolet-visible range spectrophotometry, to infrared and far infrared spectrometry, including FTIR (Fourier transform infrared spectroscopy) All these are used in analytical food and drug chemistry including in toxicology, depending on what are we looking for. Spectrophotometry is an important almost non-destructive technique used in many biochemical analyses that involve DNA, RNA, and protein isolation, enzyme kinetics and biochemical analyses. Since samples in these applications are not readily available in large quantities such as for trace evidence in forensic science, they are especially suited for small quantity non-destructive technique. Another advantage is, precious sample can be saved by utilizing a micro-volume platform where as little as 1uL of sample is required for complete analyses. Compare them with other assays, ELISA plates have the reaction products immunologically-absorbed on the solid phase, which is part of the plate, and so are not easily reusable. It is fast, simple, sensitive and specific over a range of targets and species. Basically, ELISA uses antigens from the sample that are attached to a surface. A matching antibody is then applied over the surface so it can bind to the antigen. This antibody is linked to an enzyme, and in the final step, a substance containing the enzyme's substrate is added. The subsequent reaction produces a color complex that signals a positive result. This is quite reliable, simple to read, best of all, very specific, and is not interfered by any other proteins, ligand or substances, unless of course that specific antigen is destroyed chemically or by heat treatment. But what I did not know was, ELISA has now found in application in forensic science. This is something new to me I have just learn in this current course in Forensic Toxicology from Dr. Susan Gurney at the University of Cambridge Thank you Dr. Susan for this new knowledge you gave us Lim ju boo Malaysia |
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