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must be performed carefully and deliberately to avoid disrupting vital structures. However, if the mouse begins gasping for air or stops breathing altogether and more time is needed to continue the operation, the skin should be grasped at the edges of the incision and the incision sealed. Negative intrathoracic pressure is reestablished as the mouse gasps for air. After a few minutes the mouse generally recovers, and the operation may continue.
Prior to using the thymectomized mice for experiments, it is important to confirm the absence of thymic remnants by gross and/or microscopic examination, as remnants often get pulled between the heart and the lung and might not be obvious immediately after surgery. It is not uncommon for thymic remnants to give rise to a regenerated thymus. These should not be confused with cervical lymph nodes, which may approach a size as large as the thymus. To verify the thymectomy and to distinguish thymus and lymph nodes in this region, gross and microscopic examination of the animals following their sacrifice is helpful. An alternative is to perform flow cytometry of splenic or peripheral blood cells for Thy-1+ cells (units 5.3 & 5.4), or anti-Thy-1 antibody-dependent cyto-toxicity assays of spleen cells of the putatively thymectomized animals (unit 3.4). This latter technique will only be informative for neonatal thymectomized mice, because adult thymec-tomy will leave T cells for 4 to 6 months.
Anticipated Results
After sufficient practice, a survival rate of greater than 95% can be achieved following adult splenectomy or thymectomy. For neonatal thymectomy, a survival rate of 95% can be obtained, but for many purposes, a 75% survival rate is more than adequate.
Time Considerations
Clearly, the time required for neonatal or adult thymectomy and splenectomy depends on the skill of the surgeon. With practice, dissection and removal of the organ will take 1 to 2 min. Induction of anesthesia and postoperative observation bring the total time for the opera tion to ~10 min. By having one person operate and another administer anesthesia and observe postoperatively, twenty mice can be easily thymectomized or splenectomized in 1 hr. For neonatal thymectomy, 6 to 10 mice can be done in 1 hr. The lower rate reflects the time needed to individually anesthetize the neonates (see critical parameters).
Literature Cited
Bandeira, A., Itohara, S., Bonneville, M., Burlen-Defranoux, O., Mota-Santos, T., Coutinho, A., Tonegawa, S. 1991. Extrathymic origin of intestinal epithelial lymphocytes bearing T cell antigen receptor y8. Proc. Natl. Acad. Sci. U.S.A. 88:43-47.
Miller, J.F.A.P. 1964. The thymus and the development of immunologic responsiveness. Science 144:1544-1547.
Miller, J.F.A.P. 1965. Effects of thymectomy in adult mice on immunological responsiveness. Nature 208:1337-1339.
Pantelouris, E.M., 1968. Absence of thymus in a mouse mutant. Nature 217:370-371.
Qin, S., Cobbold, S. Benjamin, R., Waldmann, H. 1989. Induction of classical transplantation tolerance in the adult. J. Exp. Med. 169:779.
Sprent, J. 1977. Migration and lifespan of lymphocytes. In B and T Cells in Immune Recognition, pp. 59-82. John Wiley & Sons, London.
Webb, S.R., Morris, C., Sprent, J.S. 1990. Ex-trathymic tolerance of mature T cells: Clonal elimination as a consequence of immunity. Cell 63:1249-1256.
Key Reference
Sjokin, K., Dalmasso, A.P., Smith, J.M., and Martinez, C. 1963. Thymectomy in newborn and adult mice. Transplantation 1:521-525.
Classic description of method of neonatal and adult thymectomy.
Contributed by J.P. Reeves and P.A. Reeves Food and Drug Administration Bethesda, Maryland
L. Thomas Chin (adult thymectomy) University of Massachusetts Worcester, Massachusetts
Survival Surgery: Removal of the Spleen or Thymus
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