Under The Microscope: Body Fluids and Substances

 

There are several ways of obtaining blood samples at autopsy and perhaps the most useful advice is what not to take. Blood should never be obtained from body cavities after evisceration, as it is almost certain to be contaminated with other body substances. The practice of scooping 'blood' - or more accurately, bloody fluid - from the paravertebral gutters or the pelvis is always unacceptable, as urine, intestinal contents, gastric contents, lymph, pleural and ascitic fluid and general tissue ooze will always find their way into such a sample and negate the reliability of analysis. When a large [1]haemothorax or [2]hemopericardium is pre- sent, it may be grudgingly acceptable to use such blood or clot, if a clean sample is taken immediately on opening the chest, before any dissection or disturbance of organs is made. This is only second best to obtaining intravascular blood - and, in the case of alcohol and other diffusible substances, the results cannot be relied upon, as the sample may be contaminated by post-mortem diffusion from the stomach, as dis- cussed in the preceding section. Carbon monoxide estimations may also be made on clean cavity blood, if nothing else is avail- able, as sometimes happens in a badly incinerated body. As it is absorbed through the lungs, it cannot diffuse from the gastrointestinal tract to give a false result. Even so, it is a rare autopsy that cannot provide a small sample of blood from some peripheral vein, if enough care and persistence is used.

To return to the general state of affairs, the most satisfactory way of obtaining a venous blood sample is venepuncture of the femoral vein by direct puncture in the groin before the autopsy begins. Practice is required as, unlike a living patient, the vein is not usually palpable. Once the lumen is entered, 25 ml can usually be drawn off without trouble, though a slow flow may be improved by massaging the leg to drive blood proximally or the leg may be raised, if rigor allows. Though less satisfactory than a leg vein, for reasons dis- cussed earlier, a neck win can also be punctured through the skin, and even after death the jugular may sometimes be both visible and palpable, especially in congestive deaths. The subclavian vein can also be tapped.

When collecting blood samples by percutaneous puncture, some pathologists prefer to use a needle designed for aspirating samples from rubber-capped vials, rather than a needle meant for clinical aspiration or injection. The same needles can be used for collecting vitreous humour. An example of this type of needle is the Becton Dickinson purple-hubbed 16 gauge one-inch or a similar American equivalent. In some jurisdictions, especially in countries where the system allows only a low autopsy rate, transcutaneous sampling is virtually the only post-mortem investigation possible and the doctors in these areas become adept at cadaver venepuncture. Many pathologists, the authors included, have been brought up to obtain blood samples during the actual performance of autopsy and this can be as satisfactory as external venepuncture, as well as saving the expense of a new syringe and needle every time. A common procedure after evisceration is to hold a container (such as the 30 ml universal) under the severed end of the subclavian vein, within the upper part of the empty thorax. The arm is then elevated and massaged if necessary, to express blood into the container. Similarly, and perhaps preferably, the femoral or external iliac vein may be transacted near the brim of the pelvis, and a container held under the cut while the leg is elevated, or massaged, or both. Either of these methods will provide ample, clean, peripheral veinous blood, through care must be taken to avoid my contamination from the body cavities. This initial collection should be transferred into one or more new rubes, with preservative added if appropriate, as the first receptacle will be soiled externally during the collection process.

An alternative method of collecting a venous sample, which always gives a copious flow of blood, is to incise the internal jugular vein during the initial stages of the autopsy. As soon as the main autopsy incision is made, the flaps of the neck skin can be dissected aside and the jugular exposed, if necessary by dividing the sternocleidomastoid muscle. As in the previous technique, a bottle can be held against the vein as it is transected and the flow collected directly into it. The blood may overflow into the pouch formed laterally by the reflected skin, which forms a reservoir where a consider- able volume may collect. If the flow is scanty, it can be improved by raising and lowering the head, which will cause blood to return from the upper venous drainage areas. Even more can be expressed by pressing on the chest, but here there is the potential disadvantage of forcing up blood through the superior vena cava from the heart, which may distort the value of alcohol and other drugs that may have diffused port-mortem from the stomach. Blood should not be taken from the heart cavities, the inferior vena cava, or the portal or hepatic veins, as these may also give concentrations that are at variance with those in the peripheral vascular system. A full 30 ml of blood should be submitted to the laboratory wherever possible, together with a 5 or 10 ml fluorided sample.

 

Prior to, or in the absence of, an autopsy, urine can be obtained by catheter or suprapubic puncture with syringe and long needle. At autopsy it is usual to wait until evisceration has been carried out before dealing with the bladder. A sample can be obtained by puncturing the fundus with syringe. and needle. Alternatively, the bladder can be stretched by pulling the fundus upwards with the fingers, then a sagittal incision made with a knife on the ventral surface, which must be free from blood soiling. The urine that wells out can be collected directly into a small container such as a 30 ml universal. If only a small amount is present, then the incision may have to be enlarged and the residual urine sucked out under direct vision, using a syringe without a needle. This can be useful for some analyses, such as those for morphine and chlorpromazine, which are concentrated by the liver and excreted into the gallbladder. Direct collection into a bottle is advised, as bile is usually too viscous to be sucked through a needle.

This is not often required for toxicological analysis, though it may be needed for microbiological and virological studies. If needed, it should be collected by lumbar or cisternal puncture, as in the living patient. It can be difficult or impossible to obtain in this way because of the lack of any intrathecal pressure. The body should be turned on to its side before the autopsy begins and flexed as much as possible by an assistant. If an infant, it should be sat up and again flexed forwards to curve the spine as much as possible. A needle on a syringe should then be passed between two lumbar or lower thoracic spines, and stopped as soon as any penetration of the theca is felt. Moderate suction should be maintained on the syringe piston to compensate for the lack of internal pressure. Alternatively, the needle should be passed into the midline just below the occiput and advanced upwards until the skull is contacted just posterior to the foramen magnum. The needle is slightly withdrawn and re-advanced until it slips through the posterior part of the atlantooccipital membrane into the basal cistern when cerebrospinal fluid may be aspirated. It is of little use taking cerebrospinal fluid from the bowl of the posterior fossa after the brain has been removed as the con- centration of many substances is different in blood compared with the fluid, so blood contamination distorts any results. Clear cerebrospinal fluid may, however, sometimes be obtained from the lateral ventricles, either by needle puncture or cutting down through the cortex.

 

Examination of Vitreous humour s is sometimes useful, especially in bodies with appreciable post-mortem decomposition, as the fluid in the eye as it was already described by me. Vitreous fluid is also used for estimating the time since death. A puncture should be made through the sclera at the outer canthus with a fine gauge needle. This should be placed as far laterally as possible, pulling the lid out, so that when released it returns to cover up the puncture mark for cosmetic reasons. The fluid should be sucked out by syringe, but it will often come only slowly because of its viscosity. If the best restoration is needed, water should be reinjected through the same needle to reinflate the globe, which tends to collapse on suction.

On the other hand, stomach contents can be collected directly into wide-mouth glass or plastic pot of at least 250 ml volume. To collect them, the exterior of the stomach should be washed clean of blood and other contamination, and pulled with attached organs to the edge of the dissecting board or sink. The greater curvature should be opened cautiously with large scissors, and the jar held underneath so that the contents flow directly into it. When most have been expressed, the greater curvature can be opened up fully and the gastric lining examined. Any further contents are scraped out and any powder, capsule or tablets picked o& and either added to the main jar or placed in a separate small container. Such focal deposits form a more concentrated sample for the laboratory. Any undissolved tablets or capsules should be carefully preserved, as the laboratory or a pharmacist may be able to identify them by their appearance and colour. After the mucosa has been examined, including that of the duodenum, the stomach wall can be dissected off. Some laboratories require this for their analysis, either added to the jar of contents or sent separately. The wishes of each individual toxicologist should be dis- covered in advance.

These are not routinely required for analysis unless some particular gastrointestinal poison is suspected, such as one of the heavy metals. Again, the toxicologist should be consulted. If required, both ends of the small gut should be ligated by string sutures at duodenum and ileum. The intestine is cut through at these points and stripped out by cutting through the mesentery in the usual way. One suture can be cut and the contents milked fr0.m one end to the other into a suitable jar, the gut then being opened and sent with the contents to the laboratory. The large intestine and contents are rarely required, as most toxicology laboratories are not keen on handling and storing large volumes of faeculent material. In heavy-metal poisoning, such as arsenic and antimony, however, some analysis may be required.

Vomit is rarely collected at autopsy, unless a large quantity has been found in the air passages. However, vomit is not infrequently collected by ambulance crews and police at the scene of the illness or agonal event, and brought with the deceased to hospital or the mortuary. If thought relevant in a case of suspected poisoning, it should be properly contained, labelled and forwarded with other samples for toxicological examination.

In solvent abuse and deaths from gaseous or volatile substances, the toxic material may be isolated from a whole lung. As soon as the thorax is opened at autopsy, a lung is mobilized and the main bronchus tied off tightly with a string ligature. The hilum is then divided and the lung laced immediately into a nylon bag, which is sealed and sent as soon as possible to the laboratory. Ordinary plastic (polythene) bags are not suitable for this task, as they are permeable to volatile substances. Nylon bags, as used by arson investigators to collect samples that may have volatile accelerants, do not suffer from this defect. In collecting blood samples for volatile substances, plastic tubes or tube syringe combinations of the 'monolete type are unsuitable, as the concentration of such compounds as toluene or other solvents will decrease considerably over a few days in storage. Vials with a rubber septum as a seal are also unsuitable, as the volatile substance can escape through the rubber. Suitable tubes are made of glass with an aluminium foil-lined cap or a polytetrafluorethylene (Teflon) liner. This type of tube is often used in laboratories for scintillation counting and can be employed for the collection of solvent samples. Such tubes should be available in any busy autopsy room where solvent abuse deaths may be handled. The tubes should be filled to the top to avoid loss of vola- tile substances into the head space and should be stored at 4"C, rather than being frozen solid.

Body tissues may be needed for some toxicological analyses. Liver is the most usual, as it concentrates many substances and their metabolites, which may then be recoverable long after the blood and urine levels have declined. Either the whole organ is saved or an aliquot of 50-100 g, according to the wishes of the articular laboratory. If only a part is retained, it should be taken from the periphery, away from major vessels and bile ducts. Brain and kidney may sometimes be required and again a 50-1OOg aliquot is usually sufficient. As with liver, the total original weight of the organ should be recorded and notified to the toxicologist. Where a toxic substance may have been injected into the subcutaneous tissue or muscle, these tissues should be excised and submitted to the laboratory. The usual method of identifying the site is from a needle puncture mark, and a zone of skin and tissue should be removed circumferentially around this or an ellipse cut away, so that the defect can be sewn up on the body. A few centimetres in diameter around the mark is usually sufficient. The depth of the sampling depends on how deeply the needle track extends and may need to go into the underlying muscle. Radiography may assist in rare cases, as in the George Markov murder in London, where a political assassination was carried out by the introduction of a tiny sphere, presumably injected from an air weapon concealed in an umbrella. The sphere, drilled out to carry a potent toxin, probably ricin, was located by X-ray at autopsy after a small skin puncture was found. Where an injection site is sampled, it is essential that a con- trol area from a remote part of the body is also sent to the laboratory. It is usual to take this from a symmetrical zone on the contralateral side, but caution must be used, as in both drug dependence and insulin usage, alternate sides may be used at frequent intervals for injection, so another more remote site might be preferable. The substances which may have been injected are numerous, but insulin, morphine, heroin, cocaine and other illicit drugs are most common. In the notorious Coppolino case in the USA, products of succinylcholine (suxarnethonium) were identified from around a needle track in the buttock of an exhumed body of a woman, leading to the conviction of her anaesthetist husband for murder.

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[1] A hemothorax (derived from hemo- [blood] + thorax [chest], plural hemothoraces) is an accumulation of blood within the pleural cavity. The symptoms of a hemothorax may include chest pain and difficulty breathing, while the clinical signs may include reduced breath sounds on the affected side and a rapid heart rate. Hemothoraces are usually caused by an injury, but they may occur spontaneously due to cancer invading the pleural cavity, as a result of a blood clotting disorder, as an unusual manifestation of endometriosis, in response to pneumothorax, or rarely in association with other conditions.

Hemothoraces are usually diagnosed using a chest X-ray, but they can be identified using other forms of imaging including ultrasound, a CT scan, or an MRI. They can be differentiated from other forms of fluid within the pleural cavity by analysing a sample of the fluid, and are defined as having a hematocrit of greater than 50% that of the person's blood. Hemothoraces may be treated by draining the blood using a chest tube. Surgery may be required if the bleeding continues. If treated, the prognosis is usually good. Complications of a hemothorax include infection within the pleural cavity and the formation of scar tissue.

 

[2] Hemopericardium refers to blood in the pericardial sac of the heart. It is clinically similar to a pericardial effusion, and, depending on the volume and rapidity with which it develops, may cause cardiac tamponade, he condition can be caused by full-thickness necrosis (death) of the myocardium (heart muscle) after myocardial infarction, chest trauma, and by over-prescription of anticoagulants. Other causes include ruptured aneurysm of sinus of Valsalva and other aneurysms of the aortic arch. Hemopericardium can be diagnosed with a chest X-ray or a chest ultrasound, and is most commonly treated with pericardiocentesis. While hemopericardium itself is not deadly, it can lead to cardiac tamponade, a condition that is fatal if left untreated. 

Acknowledgements:

www.aived.nl    AIVD – @Erik Akerboom ©

www.politie.nl  Politiekorpschef  @Janny Knol©

www.politie.nl WEB Politie - @Henk van Essen©

 

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