Introduction to DNA Profiling

Here we are, the technique considered by many to be a holy grail of forensics. But what is DNA profiling and how does it work? Well first things first, the term DNA profiling refers to the sequencing of an individuals DNA. 

When DNA profiling there are 3 possible outcomes:

1. Exclusion - the sample DNA does not match the suspects, thus ruling them out. This can be used to prove innocence or rule out suspects.

2. Inconclusive - due to damage through contamination or DNA degradation, further information must be sought, and the test must be carried out again.

3 Inclusion - The DNA is a match. However as DNA is not unique the significance of the match is calculated by quantifying the random match probability (Pm)'. The (Pm)' is a statistical test that estimates the probability of two unrelated people sharing the same DNA profile. Where markers are not linked (so inherited independently), the (Pm)' can be estimated by multiplying individual allele frequencies within a sample population. Therefore the more loci that are included in the analysis and the greater their heterozygosity the smaller the value of (Pm)' will be. A smaller (Pm)' means that the DNA being profiled is more likely to have come from the suspect. 

Despite DNA profiling's high levels of specificity there are a few problems with the technique. The first is that DNA will degrade over time, meaning it is harder to produce conclusive results with the test. The second major problem is that DNA from related people is extremely similar so has a much higher (Pm)' value. This means in cases between family members DNA is much less useful.

We'll end the introduction with a quick case study:

   1983, a girl in Narborough, Leicestershire is found dead with sigs of rape. A semen sample recovered from her body was tested and showed that the murderer was blood group A and had an enzyme profile shared by about 10% of the male population. However this was the only evidence recovered, and the investigation made no progress.

   1986, a second girl in the same area, was also found raped and murdered. The semen sample recovered from the body showed the same characteristics as the one from the case 3 years ago. The police established this connection between the 2 cases. The police believed that they were the work of the same man and that he lived in the vicinity. Their investigation led them to Richard Buckland, who was arrested. When interrogated he confessed to the second girls murder but denied having any involvement in the first. Buckland was DNA profiled, as were the semen samples. The girls had in fact been killed by the same man, but this man was not Buckland. 

   Having lost their primary suspect the police tried a different approach. They conducted the first ever mass DNA screening. All men in the vicinity (roughly 5000) had a blood sample or saliva swab taken. From these those exhibiting a matching enzyme profile and blood group where then DNA profiled. However despite all this they had nothing. There were no matches. Once again the investigation hit a wall. 

    Six months later however a woman reported that she had overheard a man claiming to have provided DNA in place of his friend, Colin Pitchfork. Colin Pitchfork was subsequently dragged in, and his DNA profile, unsurprisingly, was a match, inclusive. By 1988 he had been sentenced to life in prison. 

Colin Pitchfork on his arrest

   This case study helps show how DNA evidence alone is rarely enough to find or convict a suspect, but is very effective at excluding suspects. Thanks for reading! I'll be back soon, and we're going to be getting into some harder science! Over and Out!

Testing for Blood

You've probably all seen it in TV dramas; the crime scene looks clean as a whistle, not a trace left. That is until the CSI unit rolls in, sprays a mysterious liquid all over the place, off go the lights, out come the UV lamps and suddenly the room is covered in ethereal blue stains. Well you may be happy to learn that that's one of the few things that forensics shows largely get right.

Presumptive tests for blood reveal blood where it may not have been found with the naked eye. This could be due to it blending well with the background, seeping into cracks, or possibly just occuring in trace amounts. Luckily presumptive tests can help a forensic scientist find and identify blood at a crime scene.

Presumptive tests for blood are usually based on haemoglobin's ability to catalyse the oxidation of certain reagents. The most common reagent is Hydrogen Peroxide Solution (H202(aq)). These tests also commonly use reagents that change colour on oxidation. Phenolphthalein which goes from colourless to pink when oxidised.

To test a stain, a small circular piece of absorbant card (diameter=25mm) is folded twice to form a point. A small amount of the stain being tested is then scraped onto this point. a drop of phenophlthalein is added by a drop of hydrogen peroxide, with the bright pink colour showing a positive result. It's important to note that this test does not differentiate between human blood and other kinds of blood. 

However in some situations the colour change tests are not appropriate. Maybe the crime scene has been scrubbed clean, or you need the blood 'intact' for DNA profiling. This is were luminol comes into play. In the luminol test an alkaline solution containing both luminol and an oxidising agent (for example Hydrogen Peroxide), is sprayed onto the area being tested. Where blood is present luminol is catalytically oxidised and produces a distinct glow. 

Luminol before and after

I hope this has helped to illuminate the basics of blood identification. Until next time! Over and out!

Immunoassays in the Analysis of Drugs and Poisons

I'm back, and this time we're going to be looking at one of the most powerful techniques in the toxicologists arsenal, the immunoassay. No matter the sample size or the specificity or broadness of molecules your trying to ferret out, the immunoassay is both practical, accurate and relatively cheap.

In forensics the immunoassays are used mainly for identifying the presence of drugs in or poisons in both ante and post mortem samples. Immunoassays can be highly specific, picking out only a single compound (e.g. mephamphetamine), or a group of compounds. Immunoassays are useful because not only are they highly sensitive, they are also able to analyse large samples relatively quickly and require no preliminary extraction phase.

Immunoassay relies on the antibody-antigen reaction, which occurs in the mammalian immune system. When an antigen enters the body, an antibody specific to it i produced. The antibody binds with the antigen forming an antigen-antibody complex. In immunoassays specific antibodies are used to detect and measure the level of the analyte of interest.

In immunoassays, fixed quantities of the specific antibody to the analyte and a labelled form of that sample are added to the sample being tested. The labelled molecules and the analyte (if it's present) will competitively attempt to bind with the antibodies, in inversely proportional numbers. So you can measure the reduction in concentration of the labelled poison to work out the concentration of a drug in the sample. 

However in some types of immunoassay, it isn't possible to distinguish between the labelled poison bound to the antibody and that remaining free in solution, so physical separation of the two phases is required before measurement. These are known as heterogenous immunoassays (for example radioimmunoassay). However immunoassays where this is not necessary are called homogenous immunoassays (for example fluorescence polarisation immunoassay). These two examples are actually both very good examples of immunoassays. 

Radioimmunoassays: In radioimmunoassays, the poisonous molecules are labelled with an appropriate radioisotope (for example iodine-125) and the radioactivity is measured. The concentration of unlabelled poison molecules in the test mixture is determined by reference to a standard curve. This graph is produced by adding increasing concentrations of poison to a fixed quantity of labelled poison and antibodies. 

Fluorescence Polarised Immunoassays: A suitable fluorescent substance is used to label the analyte. It's possible to distinguish between labelled analytes bound to the antibody and analytes free in solution. This is because the former emits polarised fluorescence, but the latter generates unpolarised fluorescence.

Thanks for reading, over and out!

Jamaica ginger and 'The Jake Leg'

The Jake Leg was an epidemic found primarily during the early 20th Century in 'dry' America. America in the early 20th Century was place rife with poison and one of the places that forensic since was born. The Jake Leg however was not a symptom of poisons employed by malicious murderers or ignorant architects, rather something people induced themselves. As the astute of you may have guessed this this is symptom of bootlegged alcohol poisoning, namely a type called Ginger Jake. Ginger Jake was the dry alcohol for the poorer folk in South America, who could not afford properly, illegally distilled alcohol.

Those readers who are not experts on prohibition America (what weirdos right?) may be asking what the jake leg was. The jake leg was in fact a particularly nasty symptom of ginger jake poisoning. The jake leg was in fact a partial paralysis, which was usually severe enough to force the sufferer to walk in a foot slapping, high stepping style. 

The Jake Leg was actually caused tri-orthocresyl (TOCP). Tri-orthocresyl is an organophosphate which is used as a solvent in industry (a plasticiser), especially in aircraft oil. It causes peripheral neuropathy, degeneration of the peripheral nerves, inhibiting communication between brain and extremities. 

This effect is primarily due to the reaction between TOCP and a particular enzyme in the nerves (it acts as an enzyme inhibitor). The TOCP enzyme complex then goes through a process known as ageing. The reaction with the protein occurs rapidly (taking roughly an hour), however the neuropath takes much longer to become evident (10 days at least). This is because the enzyme which is inhibited by TOCP is essential to nerve cell function. Therefore in the long cells in the arms and legs the nerves die progressively. This entails a degeneration of the myelin sheathe and nerve working upwards towards the spinal cord. 

Dry America was a dangerous place for many reasons, with radon in water, thallium in make up and TOCP in alcohol. Toxicology is the science that put a stop to this. It doesn't just ensure that murderers are brought to justice but helps to protect everyone from exposure to some of the deadliest chemicals out there. 

That concludes todays post, until nest time! Over and out. 

Questioned Document Examination: Literary Forensics

Familiar to anyone who's watched 'Zodiac', handwriting analysis is a field which is invaluable due to the ubiquity of checks and receipts, and the scarcity of competent forgers. It has many uses as well, from confirming suicide notes to tracing who bought what drugs.

Handwriting analysis is usually made up of three stages, according to the ASTM standard guide for examination of written evidence:

1. Analysis:
   Questioned and known items are analysed and broken down to directly perceptible characteristics. These characteristics are usually:
  - Form : elements comprising the shape of the letters, such as proportions, slants, angles, retracings, connections and curves.
  - Line Quality : The pressure exerted and continuity of the script.
  - Arrangement : Any distinctive punctuation, alignment or formatting.
  - Content : Spelling, phrasing, punctuation and grammar.

2. Comparison:
   The characteristics are then compared to a known standard. A known standard, otherwise known as a writing examplar, is a piece of writing used to compare your sample against. It is either requested or collected. These are complimentary to each other as collected writing is less likely to be altered, but typically has fewer reference characteristics, while you can control the conditions perfectly and shove in as many reference points as you want to a requested piece of writing but it is much more likely to be altered. A combination of both is always a good thing.

3. Evaluation: The characteristics of the pieces of writing are evaluated and cimilarities and differences are compared. The conclusion is drawn from a combination of the frequency and uniqueness of characteristics present.

It is important to note that handwriting analysis is not a perfect science, thus in order to use it in a court of law it would always be advisable to seek peer review. With handwriting analysis there are so many variables and every style is so unique that it is difficult (if not down right impossible) to draw certain conclusions. However it is still fairly reliable, and since almost nothing is certain in forensics that's good enough to make it a useful tool in the forensic scientists arsenal.

Here's a quick example of where the FBI managed to pull a match between the sign off of the baby sitter, and the sign off on a ransom note. The minimal efforts to disguise the writing make this suspect an easy case study.

Thanks for reading everyone, any comments, critiques or suggestions, feel free to comment! If not sit tight until next time!

Animal Toxins: Snakes

Today, in a special issue of this blog, I'm going to be looking at animal poisons (toxins), specifically snakes. Despite their reputation there are only about 300 poisonous species of snakes, which is about 10% of all known species of snakes. Nevertheless, snakes do possess some of the most deadly toxins, especially cobras, vipers and sea snakes. While snake toxins are not nearly as dangerous or fast acting as some of the toxins found in plants and fungi (think ricin in castor beans, or death cap mushrooms), they can be far more deadly for two main reasons:

   1. While plants and fungi only contain a single type of toxin, snakes often utilise a mixture, making it much more difficult to treat.

   

   2. While plant or mushroom poisons have to be ingested, snake toxins are forcibly injected into your system. 

The likelihood of you dying from a snake bite is pretty much proportional to how good your country's infrastructure is. For example in North America only about 5 people per year die of snake bites, however about in India and Pakistan there are about 50,000 to 70,000 deaths per year due to snake bites.  We can tell that this is not just due to a differing number of poisonous snakes as Australia, the country with the greatest concentration of poisonous snakes has one of the lowest snake death rates.

So why is there such a discrepancy in the snake bite mortality rates. Well as with all medicine speed of response is absolutely essential, and a good infrastructure improves this. Snake bites also require adept medical professionals to treat, as I have mentioned they contain many different types of venom. North American rattlesnakes, for example, utilise at least nineteen different types of toxin. In many cases these toxins can work in a sort of perverse harmony, increasing each others' damage and rapidity of action. 

Toxins have two main ways of killing:

   1. Neurotoxins: As you may have guessed neurotoxins act on the neurons. Neurotoxins are chemical agents that affect the transmission of chemical signals between neurons. Neurotoxins affect this process in a number of different ways. They may block receptors, stopping the flow of transmission, or damaging microtubules or vesicles which carry the chemical messages. This can lead to death by asphyxiation, as chemical communication in the lungs and diaphragm is restricted, obstructing ability to breathe. 

   2.De-clotters: Ok, that 's a made-up word, but I couldn't find the technical term for this. Basically the toxin stops your blood clotting, giving you a sort of impermanent haemophilia, meaning you could actually bleed to death through the bite. The toxin may do this by inhibiting the action of coagulases. These are enzymes that convert fibrinogen into fibrin, which forms the thread of the clots. If these enzymes do not function, the blood will not clot. 

So how do we stop this? Antivenoms are the answer. Antivenoms are actually types of 'two-layer' vaccines. First the animal from which the venom comes has to be milked for its venom. The venom is then diluted to non-lethal levels and injected into an animal (a rabbit for example). The rabbit produces an immune response to the venom. These antibodies are then collected and injected into the person, defending them against the venom. 

The availability of antivenoms and the speed with which they can be received are the main factors affecting how safe you are, although not being around particularly poisonous snake infested areas cant hurt. 

Until next time people!

Gas Chromatography: Method and Applications

I've covered thin-layer chromatography previously in this blog, however chromatography is a big field, and the precision to which it can analyse samples is increasing rapidly, not to mention how widespread the apparatus required to carry it out is becoming. As such I would say it merits a lot of attention as a field of study.

Unlike in thin-layer chromatography, the mobile phase is an inert gas (for example helium), or an unreactive gas (for example nitrogen). The stationary phase is a liquid coating the walls of the column. The stationary phase is chosen to be as close to the polarity of the solute as possible.

The gas chromatograph uses a column (flow through narrow tube). The different constituent chemicals from the analyte are carried up the column by the mobile phase at different rates. The rate is dependant on the chemical and physical properties of the chemicals with the stationary phase. This causes each constituent to exit the column at a different time.

You may be wondering why such complex, and often expensive techniques need to be employed, well fear not I hear your worries and I'm going to illustrate the value of gas chromatography, thin-layer chromatography and mass spectrometry with a couple of case studies.

1. FBI's analysis of EDTA in blood at O.J. Simpson trial:
    Agent Martz and the FBI forensics team used a combination of the aforementioned techniques to detect the presence (or in this case absence) of EDTA in a blood sample from socks and a watch. EDTA is a preservative for blood. If found in high levels it would indicate that the blood samples had been tampered with. The high level of precision, and ability to remove background peaks, of the mass spectrometer meant that this case was where it really established its role as a reliable form of evidence.

2. Analysis of Fire Debris for ignitable liquids:
    In January 2002 there is a fire. One person dies in the fire, and the police decide to bring in a forensics team to investigate. The result is a piece of vital evidence, showing the fire could not have been an accident. The forensics team, using Gas-Liquid Chromatography and Mass Spectrometry find gasoline present on weathered debris. The presence of ignitable liquids (where there should be none) is fairly damning.

Anyway thank you for reading, until next time people.