The following guidelines have been approved by the University Institutional Animal Care and Use Committee to insure that animals used for the production of hybridomas for the production of monoclonal antibodies experience minimal pain and distress .

• Immunization volumes should not exceed 0.2 ml when administered subcutaneously or 0.5 ml when administered by intraperitoneal injection. In vitro immunization procedures have been reported to be beneficial when antigen is in limited supply and previous attempts at in vivo immunization have failed. Other advantages of in vitro immunization are the requirement for fewer animals and the immunization period is reduced to 4-5 days^1,2,3. The investigator should review the IACUC Recommendations for Use and Alternatives to Freund's Complete Adjuvant before selecting an adjuvant for use.
• It is recommended that hybridomas and the monoclonal produced be tested for the presence of adventitious viral and mycoplasma agents prior to use in animals. Viral contamination is common in murine leukemia and transplantable tumor specimens. Inoculation may result in infection of the animal and spread of disease through the vivarium. Also, lymphocytic choriomeningitis virus has been isolated from tumor cell lines which is capable of producing illness in humans⁴.

Adapted from "Institutional Policies and Guidelines on Adjuvants and Antibody Production"; Jackson LR, Fox JG. ILAR Journal Vol 37, #3, p141-52, 1995.

1-Borrebaeck CAK, Strategy for the production of human monoclonal antibodies using in vitro activated B cells. J. Immunol. Methods 123:157-65, 1989.

2-Hengartner H, Luzzati AL, Schreier M, Fusion of in vitro immunized lymphoid cells with X63Ag8. Curr. Top. Microbiol. Immunol. 81:92-99, 1978.

3-Bunse R, Heinz H-P, Characterization of a monoclonal antibody to the capsule of Haemophilus influenzae type b, generated by in vitro immunization. J. Immunol. Methods 177:88-99, 1994.

4-Dykewicz CA et al. Lymphocytic choriomeningitis outbreak associated with nude mice in a research institute. J. Am. Med. Assoc. 267:1349-53, 1992.