Yi J Sample | collectionoe Handbook of laboratory diagnosis in swine SAMPLING AT NECROPSY Several protocols to perform a necropsy in pigs are available. The veterinary surgeon may choose any of these protocols, but itis recom- mended to follow the same procedure to per- form a systematic, complete and organised necropsy. This chapter reviews the methods and types of samples that can be used during the necropsy. A properly performed necropsy can yield much valuable information about a disease. Gross pathology alone will provide a diagnosis in some cases, but additional testing is often required to obtain a definitive diagnosis. The range of techniques in use is expanding, but iraditional methods are still the first line of investigation. Table 1 gives a list of potential samples to be collected at necropsy. Selection of animals The best way to ensure an accurate diagnosis, is to have fresh and well-preserved samples. It is therefore advised to perform necropsies on recently slaughtered animals, and 3-4 pigs may thus be selected from the population of affected animals. When possible, select those animals in the acute phase of the disease (first 24-48 hours) that represent a pattern of clinical signs that is similar to that of the other members of the group. To avoid any interfer- ence with further laboratory analyses, avoid those that have already been treated (at least systemically). There are several approved protocols to perform ethic euthanasia in pigs; however, intravenous barbiturates are recommended because their price is low and they allow organ preservation. Blood sampling Blood samples should be collected in vivo (figs. 1 and 2) or just after euthanasia, when the heart is still beating. If several hours have passed after death, heart blood clots may be collected and used for microbiological, virologi- cal, PCR or serological analyses. Different in vivo techniques using various sites have been described: anterior vena cava (younger animals, from birth to 2 months of age approximately), jugular vein (fatteners and finishers) and tail or ear veins (adults). Blood samples for haematology and biochemistry should be collected into tubes containing sodi- um EDTA or lithium heparin. Blood collected into tubes without any anticoagulants is also suitable for most serological and biochemical tests (fig. 3). Histopathology Tissue samples for histopathological exami- nation, IHC analysis and ISH should be immersed in 10% neutral buffered forma- lin and stored at room temperature. The tissue:formalin volume ratio should be no less than 1:10. The sample jars must be clearly labelled with the animal's identification. All the samples from the same animal can be included in the same jar. Use as many jars as the number of necropsied pigs. Tissue samples should be no thicker than 0.5 cm; otherwise the lack of penetration of formalin may favour autolysis (fig. 4). The exception is the brain and eye globe, which are fixed intact. As for the intestine, itis recommended to collect several portions and the samples should be opened longitudinally before being immersed in formalin. The samples should be taken from the bor- der of the lesions or, alternatively, include affect- ed and non-affected areas of the same organ. Microbiology/virology/ molecular techniques Specimens of blood, urine, saliva, milk, cerebrospinal fluid or tissues should be col- lected as aseptically as possible for culture, virological analysis or PCR tests (figs. 5 and 6).jgure 1. Biood collection n vivo from the cranial cava vein Figure 2. Blood collection in vivo from the jugular vein. Figure 3. Blood collection tubes for haematology and biochemistry with an anticoagulant (EDTA - pink, heparin - green). Tubes without any anticoagulants (ed) are used to obtain serum in order to perform serological or biochemistry tests. Figure 4. Tissue collection for histopathology. Samples should nat be thicker than 0.5 cm to facilitate fixation; the intestine must be longitudinally opened