BACKGROUND
Guinea pigs are the animals of choice for skin sensitization tests. Guinea Pig Maximisation Test (GPMT) is among the mostly used methods for testing of chemicals topically.1
The basic principle involved in this procedure was skin sensitization is an immunological process in which the test animal acquires a specific allergic sensitivity to test substance on repeated exposure and the response is manifested as increased erythema.2
At first test animals are injected the test substances by intradermal or epidermal route (induction exposure). The animals are then exposed to a test dose. The extent and degree of skin reaction in test animals is compared with control animals which undergo sham treatment during the period.
REQUIREMENTS
Animals: Guinea pigs, both sexes
Chemicals: Frauds complete adjuvant (FCA)
Miscellaneous: Physiological saline, Syringe, Shaving kit, Whatman filter paper no. 1
PROCEDURE
Weigh and select healthy young adult animals of both sexes for the study. Female animals should be nulliparous and non-pregnant. Take 10 animals for test group and 5 animals for control group. Animals should be weighed before and after the test.
Procedure involves testing the drug in two phases, i.e., induction phase and challenge phase.
Induction phase
For test group
Prepare three pairs of intradermal injections. First injection contains 1:1 (v/v) mixture of Frauds complete adjuvant (FCA)/water or physiological saline.
Injection 2 contains a test substance mixed in appropriate vehicle at selected concentration and 3rd injection contains test substance at required concentration formulated in 1:1 (v/v) mixture of FCA/water or physiological salt solution.
(For injection 3 if test substance is water soluble dissolve the same in aqueous phase first than mix with FCA. If test compound is lipid soluble then suspend the drug in FCA and then in aqueous phase)
Shave the shoulder portion and caudal region of the animal either by clipping, shaving or chemical depilation. Inject the first and second injection through intradermal route at a dose of 0.1 ml on both the sides of shoulder (nearest the head) and third injection on the caudal area.
For control group
Inject three pairs of 0.1 ml intradermal injections similarly at the same sites as in treatment group without test drug in preparations. Observe the treated sites of each animal and score at 24 and 48 hours after injection.
After 7 days of primary induction phase, the test area is once again shaved cleanly and the test drug is administered topically by applying test drug saturated 2 cm x4 cm section Whatman filter paper no. 1 over the sites treated previously for 48 hours.
The animals in the control group are treated in a similar manner but without test drug. After 48 hours the treated sites are examined and scored.
Challenge phase
After 14 days of topical induction, the flanks of animals in both the groups are shaved.
A patch of 2 cm × 2 cm section of Whatman filter paper no. 1 loaded with highest non-irritating concentration of the test drug is placed on the left flank and only vehicle loaded paper is to be placed on right flank (in case of solids they should be grounded finely and mixed in vehicle; in case of liquid it should be used in undiluted form).
These patches are held in contact with flanks by occlusive dressing for 24 hours. After 24 and 48 hours the patch will be removed, examined and scored.
Sensitization potential is determined by positive response showed by percentage of animals involved in the study. A score 1 or greater for redness is considered as a positive reaction of test substance.
Table 1: Magnusson and Kligman grading scale for the evaluation of challenge patch test reactions.2
| Score | Evaluation |
| 0 | No visible change |
| 1 | Discrete or patchy erythema |
| 2 | Moderate and confluent erythema |
| 3 | Intense erythema and swelling |
Observation table:
| Groups | Scoring at induction phase | Scoring at challenging phase | ||
| At 24 hours | At 48 hours | At 24 hours | At 48 hours | |
| Control | ||||
| Test | ||||
CONCLUSION
Comparison of scoring between study groups estimates the sensitization potential of test drug.
REFERENCES
- OECD Guideline For Testing Of Chemicals, 1992. Available at: https://ntp.niehs.nih.gov/iccvam/suppdocs/feddocs/oecd/oecdtg406.pdf.
- Magnusson B, Fregert S, Wahlberg J. Determination of skin sensitization potential of chemicals. Predictive testing in guinea pigs. Arbete och Hälsa, 1979;26.
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