Guidelines for Adjuvant Use

Guidelines for
Adjuvant Use

There are four basic concerns that must be considered with adjuvant use:

the species in which we use these substances
the anatomical site of injection
the route of injection
the dose of the agent per site

A fifth question, and an important one from the standpoint of regulatory reporting requirements, is whether or not we consider this a painful procedure that is not relieved by analgesics. If we do indeed consider it thusly, the investigator must justify the procedure in writing, and the justification must be submitted to the USDA along with our Annual Report of Research Facility where the procedure would be listed under Column E (unrelieved pain)

The following species and routes are typically used with Freund's Complete Adjuvant (FCA), Freund's Incomplete Adjuvant (FIA), and a variety of other adjuvants.

Mice:

Intraperitoneal
Subcutaneous

Rabbits:

Intrasplenic
Intranodal
Itramuscular
Subcutaneous
Intraperitoneal
Intadermal

Guinea Pigs:

Intramuscular
Subcutaneous
Intadermal

Chickens / Turkeys:

Intramuscular
Intradermal
Subcutaneous
Bursa of Fabricus
Intraperitoneal:

Sheep / Goats:

Intramuscular
Intradermal
Subcutaneous

Problems may arise for one or more of the following reasons:

Failure to tap off excess ascites fluid from the abdomen resulting in over-distension and distress.
Excessive dose in a single subcutaneous or intramuscular site in any species, resulting in severe abscessation, necrosis, and fistulous tracts.
Too many injection sites too close together, producing severe necrotizing dermatitis.
Bilateral simultaneous intramuscular injections in thigh muscles, resulting in lameness of the animal.
Footpad injections in any species. These result in severe inflammation, lameness, and self mutilation. It is universally held today that this practice has no basis in scientific necessity.
Hypersensitivity pneumonitis and embolic pneumonia associated with the use of FCA and FIA which may occur even when these substances are not administered intravenously. These materials should never be deliberately administered intravenously.
Sterile peritonitis and abdominal adhesions associated with intraperitoneal administration of adjuvants.

The IACUC's position on the use of adjuvants is that properly administered adjuvant followed by proper care and observation is temporarily painful but not debilitating. The use of adjuvants will be reported as a USDA Column E listing. Conversely, some practices as indicated above, are not humane and should be generally prohibited. In spite of commercial claims that some adjuvants are not inflammatory, the limited objective data available tends not to support these claims. Hence, a single, uniform practice is recommended for all adjuvants, unless there is substantive data to indicate otherwise.

SUGGESTED ROUTES AND DOSES: In all instances, adjuvant injections may be repeated in 14-30 days unless complications develop. NOTE: the following recommended volumes are for adjuvant only. Total injected volume should not exceed twice the recommended amount of adjuvant.

MICE:

Intraperitoneal. Maximum of 0.1 - 0.3 ml of adjuvant or pristane. Mice must be observed daily and tapped whenever ascites is apparent, usually every two days. Whenever mice appear to be distressed or not eating they should be humanely euthanized.
Subcutaneous. 0.01 - 0.2 ml, back of neck or inguinal area.

RABBITS: Rabbits must be carefully restrained or lightly anesthetized with ketamine if multiple injections are to be administered.

Intramuscular. Multiple sites, lateral to the vertebral column on the back; maximum of 0.3 ml of adjuvant per site. Maximum of four (4) sites. If the thigh muscle is used, only one leg, maximum of 0.3 ml may be used.
Subcutaneous. Multiple sites, maximum of 0.3 ml of adjuvant per site. Maximum of six (6) sites. Usually administered lateral to the vertebral column.
Intradermal. Multiple sites, usually lateral to the vertebral column; maximum of 0.05 ml of adjuvant per site. Injection sites should be at least 1.0 cm apart. Maximum of ten (10) sites per challenge date. Use alternate route until epidermis is healed. Intradermal injections may be repeated once skin is healed.
Intraperitoneal. Maximum of 0.5 ml of adjuvant per injection interval. Rabbit to be observed daily; euthanize humanely if rabbit exhibits distress.

GUINEA PIGS:

Intramuscular. Only a single site (thigh) is recommended. Dose not to exceed 0.2 ml adjuvant.
Subcutaneous. Multiple sites along the back. Maximum of 0.2 ml adjuvant/site and 4 sites.
Intradermal. Multiple sites along the back. Maximum of 0.03 ml adjuvant/site and 6 sites.

SHEEP AND GOATS:

Subcutaneous. Multiple sites lateral to the vertebral column, over the lateral thigh, or lateral thorax. Not to exceed 0.5 ml of adjuvant per site. Maximum of ten (10) sites.
Intramuscular. Multiple sites lateral to the vertebral column, not to exceed 0.3 ml of adjuvant per site. Maximum of ten (10) sites. Thigh muscle injections may be made in only one leg in two single sites, not to exceed 0.5 ml of adjuvant per injection.
Intradermal. Similar to subcutaneous route, but amount not to exceed 0.05 ml of adjuvant per site per challenge date.

CHICKENS AND TURKEYS:

Intramuscular. (Breast muscle). 0.2 ml of adjuvant per site. Maximum of two (2) sites per challenge date. Euthanize humanely if animal exhibits distress.
Intradermal. 0.05 ml of adjuvant per site. Maximum of ten (10) sites per challenge date. Do not repeat until all lesions have healed. Use alternate method (IM) if boost is necessary.
Subcutaneous. 0.2 ml of adjuvant per site. Maximum of four (4) sites per challenge date.
Bursa of Fabricus. Same as mouse IP.
IP - Maximum of 0.5 ml of adjuvants, single injection. Observe daily. Tap when distended. Euthanize if bird exhibits distress.

Important Notes:

FCA may only be used with the initial injection in any of the above schemes.
Footpad injections should not be used in any species.
All species require daily observation and where infected abscesses develop, antibiotic therapy should be instituted. FIA & FCA should not be used intravenously in any species.
When distress or pain become apparent from any immunization method, analgesics MUST be administered or the animal euthanized.

Exceptions (e.g., route of administration, site of administration, species, volume administered) to this guideline will be assessed based on scientific justification on an individual basis by the IACUC and veterinary staff.