Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
Animals female non-pregnant nulliparous
- Full barrier in an air-conditioned room
- Temperature: 22 ± 3 °C
- Relative humidity: 55 ± 10%
- Artificial light, sequence being 12 hours light, 12 hours dark
- Air change: at least 10 x / hour

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Remarks:
AOO (3+1 (v/v) Acetone/Olive Oil
Concentration:
25%, 12,5% and 6,25%
No. of animals per dose:
5 mice per test group
Details on study design:
Topical Application
Each mouse was treated by topical application of 25 μL of the selected solution to the entire dorsal surface of each ear.
Topical applications were performed once daily over three consecutive days.
Administration of 3H-methyl thymidine
Five days after the first topical application all mice were dosed with 20 μCi 3H-methyl thymidine by intravenous injection (tail vein) of 250μL of 3H-methyl thymidine, diluted to a working concentration of 80μCi/mL.
Preparation of cell suspension
Approximately 5 hours after 3H-methyl thymidine-injection all mice were sacrificed by cervical dislocation. The draining “auricular lymph nodes” were excised, individually pooled for each animal (2 lymph nodes per animal) and collected in phosphate buffered saline (PBS). A single cell suspension of pooled lymph node cells was prepared by gentle mechanical disaggregation through polyamide gauze (200 mesh size). After washing the gauze with PBS the cell suspension was pelleted in a centrifuge. The supernatant was discarded and the pellets were resuspended with PBS. This washing procedure was repeated.
After the final wash each pellet was resuspended in approx. 1 mL 5% TCA at approx. 4 °C for approximately 18 hours for precipitation of macromolecules. Each precipitate was once washed again, resuspended in 1 mL 5% TCA and 7 mL scintillation fluid was added. Then this solution was transferred into scintillation vials and stored at room temperature overnight.
Determination of incorporated 3H-methyl thymidine
The 3H-methyl thymidine – incorporation was measured in a ß-counter and expressed as the number of disintegrations per minute (DPM). Similarly, background 3H-methyl thymidine levels were also measured (5% TCA). Determination of radioactivity was performed individually for each animal.
Positive control substance(s):
other: P-Phenylenediamine
Statistics:
Could not be calculated as the stimulation indices of all concentrations were below 3

Results and discussion

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: The stimulation index at a concentration of 6.25% was 0.8 The stimulation index at a concentration of 12.5% was 1.2 The stimulation index at a concentration of 25% was 0.9
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: DPM Node (Negative Control) 579,4 +/-86,8

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information
Conclusions:
According to OECD 429 and the criteria given in Annex I of Regulation (EC) 1272/2008, the test item ALUFLUX or Aluminium dross or gray Aluminium dross, as described in this report is expected to have no sensitising properties and therefore, should not be regarded as a dermal sensitiser.
Executive summary:

Three groups, each of 5 female mice were treated with 25μl of the test item suspended in acetone/olive oil carrier (3:1) at concentrations 25, 12.5 and 6.25% w/v. A fourth group received only the carrier. The item was applied topically on the entire dorsal surface of each ear on Day 1, 2 and 3. On Day 6 mice were dosed with 20μCi3H-methyl thymidine and 5 h later they were sacrificed. The thymidine incorporation in the excised and pooled (per group) lymph nodes was measured. The mean stimulation index (SI) (pooled radioactive incorporation in group/pooled radioactive incorporation control) of each group was calculated. A historical positive group (phenylenediamine) was included. Details of housing and feeding conditions are given. Body weights were recorded individually and were within normal fluctuations. All the animals survived throughout the test without clinical signs. None of the three concentrations reached an SI >3, consequently aluminium dross is not expected to have sensitizing effects.