IACUC Guideline #12
Use of Freunds Complete Adjuvant
The USDA has declared that the use of Freunds Complete Adjuvant (FCA) is potentially painful or distressful. Improper use of FCA may result in undesirable and potentially painful side effects, such as excessive inflammation and swelling, abscess formation, ulceration and tissue necrosis.
The KSU IACUC is responsible for ensuring that investigators consider alternatives to any procedures that cause more than slight momentary pain. A written narrative description of the methods used and sources used to search for an alternative is required for potentially painful or distressful procedures.
The use of FCA is an important biological resource for investigators and is regarded scientifically as an effective means of potentiating immune responses in laboratory animals. Researchers have been reluctant to discontinue use of FCA because it is used as a gold standard due to its well-known effectiveness with a wide variety of antigens.
The KSU Institutional Animal Care and Use Committee (IACUC) and the KSU Office for Laboratory Animal Resources (OLAR) have determined that the potential for pain or distress may be attenuated by using appropriate procedures known to minimize the adverse effects and discomfort associated with the use of this agent.
The IACUC will therefore approve the use of FCA without an alternative search, if the guidelines described below are strictly followed. Departure from these guidelines will necessitate a written narrative description of the methods and sources used to search for alternatives to the use of FCA. The following guidelines are intended to
eliminate or reduce to a minimum animal discomfort associated with the use of FCA in research. Departures from these guidelines will require adequate justification to the IACUC.
- Before using FCA, the use of other adjuvants must be considered (i.e., RIBI, Titremax, etc.). FCA must be used only when absolutely necessary (i.e., small amounts of antigen available, weak antigens, etc.). AT this time there are no comprehensive or independent studies comparing a wide variety of these alternative adjuvants to each other.
There are some limited studies that make some comparisons. These studies never look at all of these variables; they generally only compare a couple of adjuvants to each other and often not published in peer-reviewed journals. No one adjuvant works best with all antigens, animal species, or experimental conditions. Each adjuvant has advantages
and disadvantages, and the antigen�s properties must also be taken into account to select the adjuvant most likely to give the best results. The IACUC recognizes that the principal Investigator is best qualified to select the appropriate adjuvant to be used. Information about alternatives to FCA is available at the IACUC Office, or at the
USDA website "Information Resources for Adjuvants and Antibody Production: Comparisons and alternative Technologies" (www.nal.usda.gov/awic/pubs/antibody).
- FCA should be used only for the first priming) antigenic dose. Since careful initial use of FCA does not result in pain or discomfort to the animal, the consideration of alternatives to potentially painful procedures (question #5; Standard Protocol for the Production of Polyclonal Antibodies in Rabbits) is not required.
- Intravenous, intraperitoneal or footpad injection of FCA will require the consideration of alternatives to potentially painful procedures and completion of question #5 (Standard Protocol for the Production of Polyclonal Antibodies in Rabbits). Use of two or more does of FCA is rarely warranted, but will also require consideration of
alternatives and justification for its multiple use. Each animal use protocol proposing IV, IP, footpad or multiple use of FCA will be considered and reviewed by the IACUC on a case by case basis.
- Many of the undesirable side effects of FCA can be reduced or eliminated through refinements in its use. Injection sites should be cleaned and free from debris and contamination. This is probably the major cause of abscess formation in animals, and is why some researchers have no problem with FCA and others have large problems.
Subcutaneous, intramuscular or intradermal routes, with separation between inoculation sites adequate to avoid coalescence, and the use of a small amount of inoculum per site are preferred.
- The inoculation should be kept to the minimum volume practical and should not exceed the following guidelines:
- For subcutaneous inoculation, no more than 0.25-0.5 ml per injection site. Number of injection sites should not exceed 5, for a total volume per injection of 1 to 2 ml.
- For intramuscular inoculation, no more than 0.25-0.5 ml per injection site. Number of injection sites should not exceed 5, for a total volume per injection of 1 to 2 ml.
- For intradermal inoculation, no more than 0.1-0.2 ml per injection site. Number of injection sites should not exceed 5, for a total volume per injection of 1 to 2 ml.
LAST REVIEWED AND ADOPTED BY THE IACUC: July 21, 2011