This test is found in an informative annex of the document.
The hamster is the preferred model.
There is an exemption statement that test articles are excluded when the pH is less than or equal to 2 or greater than or equal to 11.5 or if it is already known as an irritant in other tissues.
Liquids or extracts should be tested by soaking absorptive pellets and applying them directly to the cheek pouches. Solids test articles should be applied as pellets to the cheek pouches. Solids, and extracts thereof, should be prepared according to the appropriate associated annexes. If extracts are used, they should be prepared using both polar and nonpolar vehicles.
The animals used should be Syrian Hamsters of either sex of a single, outbred strain.
A total of six animals should be used. Three animals for each of the test and control (material or solvent) articles, separate groups for each extract vehicle.
In seven days prior to the study, an acclimation process is performed where the animals are fitted with collars which are designed to prevent the ejection of the test or control pellets following dosing. Animals are weighed every day for 7 days. If they lose weight they are excluded from the study.
The test article is dosed in one cheek pouch of each animal and the other is an untreated control. If the test article is a liquid or an extract has been prepared, a cotton-wool pellet can be soaked with the dose solution or the liquid can be directly instilled. If an extract of the test article is prepared, a corresponding control extract should be tested. Following dosing, the collar is put back on the animal. The procedure should be repeated hourly for 4 hours for an acute exposure and repeat exposures should mimic clinical exposure. Each cheek pouch is examined macroscopically following removal of pellets and before next dose
Macroscopic grading is assessed on a 5 point scale ranging from no(0), very slight(1), well-defined(2), moderate(3), to severe(4) for erythema. 24 hours after the last dose, the animals are scored macroscopically, sacrificed and then the cheek pouches are harvested and fixed to be processed histologically.
Macroscopic evaluation:
Test cheek pouches are compared to the contralateral control. An average score is determined for each animal with a consideration of procedural irritation.
Microscopic evaluation:
An overall average per group score is determined with the maximum score being 16. Four microscopic categories are individually scored with ranges from 0 to 4 for epithilium, leukocyte infiltration, vascular congestion and edema. The final evaluation is a 5 category scale which spans from not an irritant to a severe irritant.
The hamster is the preferred model.
There is an exemption statement that test articles are excluded when the pH is less than or equal to 2 or greater than or equal to 11.5 or if it is already known as an irritant in other tissues.
Liquids or extracts should be tested by soaking absorptive pellets and applying them directly to the cheek pouches. Solids test articles should be applied as pellets to the cheek pouches. Solids, and extracts thereof, should be prepared according to the appropriate associated annexes. If extracts are used, they should be prepared using both polar and nonpolar vehicles.
The animals used should be Syrian Hamsters of either sex of a single, outbred strain.
A total of six animals should be used. Three animals for each of the test and control (material or solvent) articles, separate groups for each extract vehicle.
In seven days prior to the study, an acclimation process is performed where the animals are fitted with collars which are designed to prevent the ejection of the test or control pellets following dosing. Animals are weighed every day for 7 days. If they lose weight they are excluded from the study.
The test article is dosed in one cheek pouch of each animal and the other is an untreated control. If the test article is a liquid or an extract has been prepared, a cotton-wool pellet can be soaked with the dose solution or the liquid can be directly instilled. If an extract of the test article is prepared, a corresponding control extract should be tested. Following dosing, the collar is put back on the animal. The procedure should be repeated hourly for 4 hours for an acute exposure and repeat exposures should mimic clinical exposure. Each cheek pouch is examined macroscopically following removal of pellets and before next dose
Macroscopic grading is assessed on a 5 point scale ranging from no(0), very slight(1), well-defined(2), moderate(3), to severe(4) for erythema. 24 hours after the last dose, the animals are scored macroscopically, sacrificed and then the cheek pouches are harvested and fixed to be processed histologically.
Macroscopic evaluation:
Test cheek pouches are compared to the contralateral control. An average score is determined for each animal with a consideration of procedural irritation.
Microscopic evaluation:
An overall average per group score is determined with the maximum score being 16. Four microscopic categories are individually scored with ranges from 0 to 4 for epithilium, leukocyte infiltration, vascular congestion and edema. The final evaluation is a 5 category scale which spans from not an irritant to a severe irritant.
