Related Experiment Videos
The assessment of micronucleated polychromatic erythrocytes in rat bone marrow. Technical and statistical
1ICI Pharmaceuticals Division, Macclesfield, Chesire, Great Britain.
Abstract:
Traditionally, the mouse is the species of choice for the rodent bone marrow micronucleus assay (MN). However, the rat is used for most other toxicological studies. The suitability of the rat as a test species for the MN was therefore investigated. In this paper, the methodological aspects of the assay have been considered. The distribution and incidence of micronucleated polychromatic erythrocytes (MPEs) on bone marrow slides prepared by two techniques, the conventional smear and the paint-brush technique, were assessed in control and cyclophosphamide-dosed male and females rats. MPEs were shown to be homogeneously distributed when assessed over a large number of PEs on slides prepared by both techniques, but when viewed over a few hundred PEs (less than 500 PEs), the incidence of MPEs on the same slides was seen to vary considerably (0-10 MPEs/500 PEs). Variability was within acceptable limits when at least 1000 PEs/animal were analysed. The spontaneous incidence of MPEs in the AP rat is low (0-2 MPEs/1000 PEs). Cyclophosphamide increased the incidence markedly and there was a wide inter-animal variability in the response (10-40 MPEs/1000 PEs). The paint-brush technique is considered technically simpler and recommended over the smear technique. This study shows that MPEs can be accurately scored in the bone-marrow of the rate provided due consideration is given to staining and sample size of PEs analysed per animal.
Insights
The rat is a suitable species for the rodent bone marrow micronucleus assay (MN). Methodological considerations, including sample size and staining, are crucial for accurate micronucleated polychromatic erythrocyte (MPE) scoring.
Area of Science:
- Toxicology
- Genetics
- Hematology
Background:
- The mouse is the traditional species for the rodent bone marrow micronucleus assay (MN).
- Rats are commonly used in other toxicological studies, necessitating an evaluation of their suitability for the MN assay.
- Investigating the rat as a test species for the MN assay is important for harmonizing toxicological testing.
Purpose of the Study:
- To assess the suitability of the rat as a test species for the rodent bone marrow micronucleus assay (MN).
- To investigate methodological aspects of the MN assay in rats, focusing on the distribution and incidence of micronucleated polychromatic erythrocytes (MPEs).
Main Methods:
- Comparison of two slide preparation techniques: conventional smear and paint-brush technique.
- Assessment of MPE distribution and incidence in control and cyclophosphamide-dosed male and female rats.
- Analysis of MPEs in relation to the number of polychromatic erythrocytes (PEs) analyzed per animal.
Main Results:
- MPEs showed homogeneous distribution over large numbers of PEs, but incidence varied considerably when fewer than 500 PEs were analyzed.
- Variability in MPE incidence was acceptable when at least 1000 PEs per animal were analyzed.
- The spontaneous MPE incidence in Sprague-Dawley rats is low (0-2 MPEs/1000 PEs), while cyclophosphamide significantly increased incidence with considerable inter-animal variability (10-40 MPEs/1000 PEs).
Conclusions:
- The rat is a suitable species for the bone marrow micronucleus assay, provided careful consideration is given to staining and sample size.
- The paint-brush technique is recommended over the smear technique due to its technical simplicity.
- Accurate scoring of MPEs in rat bone marrow is achievable with appropriate methodological controls.