Materials / Reagents Required :

            10 % Neutral Buffered Formalin (NBF), Normal saline (10 litre), 70 % Ethanol (1 litre), different sizes of scissors, forceps, scalpel, spatula, bone cutter, cardboard paper, etc.

Safety Precautions :

            Protective equipments including laboratory coats, gloves, apron, etc., should be worn during euthanasia, necropsy as well as during the preparation of the solutions.

Procedure :

• The thymus is a bi-lobed organ located in the mediastinum, anterior to the major vessels of the heart. The spleen has a smooth ivory or pink surface with a distinct lobular segmentation. Its apex is near the larynx cranially, while the curved base is attached to the cranial and ventral pericardium.
• The two independent lobes, attached to each other only by thin connective tissue, and surrounded by a thin fibrous capsular membrane, consist of smaller lobules which are also separated from each other by thin fibrous tissue septa. Blood vessels enter the lobules at the cortico-medullar junction, and extend radially into the cortex.
• Nerves course along the blood vasculature. The Lymph nodes are connected by lymph vessels and consists of cortex and medulla. The arterial blood supply, entering the node at the medulla, ends in the paracortex as arteriolar capillaries.
• These feed venules that are lined by high endothelial cells. The spleen consists of two main compartments, the red pulp and the white pulp. It is an elongate flattened organ with round ends, which is attached to the greater curvature of the stomach. A transverse section of spleen appears like a flattened isoceles triangle.
• The spleen is approximately 3-5 cm in length and about 8-10 mm in width in adult rats. The spleen roughly weighs 750-1400 mg. The size of the spleen in females is lesser than that of the male rats. Through the omentum attached to the hilus of the spleen, blood vessels  enter and leave the spleen.
• Observe that the dorsal parietal surface of the spleen faces the diaphragm and liver, while the ventral part touches the lateral abdominal wall. The spleen is supplied by the splenic artery, a branch of the coeliac artery. The lymph node is covered by a thin fibrous capsule of fibrous connective tissue. The afferent lymphatics penetrate the capsule to empty into the subcapsular sinus.

Collection of Immune organs:

• After induction of anaesthesia/euthanasia, exsanguination shall be accomplished by severing the femoral arteries, abdominal aorta or by incising the skin at the neck region. Normal saline shall be kept ready during the necropsy to keep the lungs moist.
• The fur of the animal shall be moistened with alcohol to prevent the fur from getting adhered into the tissue samples. Skin of the abdomen (in the shape of V) along with mammary gland shall be removed first. The reflected flap of the skin of abdomen is cut to expose the sternum.
• The gastro-intestinal (GI) tract along with mesenteric lymph nodes, pancreas and spleen shall be removed. The spleen present above the liver shall be separated. Place the collected spleen on a petri dish and observe the features of the spleen.
• The mesenteric lymph nodes are then separated from the mesentery. Place the collected mesenteric lymph node on a petri dish and observe the features of it.  The diaphragm beneath the sternum is cut open and the sternum is removed by two parallel cuts from the xiphoid end up to the first rib.
• The access to the thymus is facilitated by enlarging the chest open using forceps. The thymus present above the heart shall be separated, placed over the cardboard and then fixed. Place the collection on a petri dish and observe the features of the thymus.