Introduction: In vivo genotoxicity assays, especially the micronucleus (MN) and chromosomal aberration (CA) assays, are essential tools in the safety evaluation of pharmaceuticals. This quiz set focuses on practical, regulatory and mechanistic aspects of rodent-based MN and bone marrow CA tests used in M.Pharm pharmacology and toxicology training. Questions cover biological basis, specimen collection, slide preparation, staining, scoring criteria, controls, interpretation of clastogenic versus aneugenic effects, OECD guidelines, and common pitfalls. Use this set to reinforce experimental planning, data interpretation and regulatory expectations for in vivo genotoxicity studies.
Q1. Which OECD guideline specifically describes the mammalian erythrocyte micronucleus test used for in vivo genotoxicity assessment?
- OECD TG 471
- OECD TG 474
- OECD TG 476
- OECD TG 475
Correct Answer: OECD TG 474
Q2. In a standard rodent bone marrow micronucleus assay, which cell population is most commonly scored for micronuclei?
- Normochromatic erythrocytes (NCEs)
- Polychromatic erythrocytes (PCEs) / immature erythrocytes
- Granulocytes
- Lymphocytes
Correct Answer: Polychromatic erythrocytes (PCEs) / immature erythrocytes
Q3. Which chemical is routinely used as a positive control for in vivo rodent micronucleus and chromosomal aberration tests?
- Sodium chloride
- Cyclophosphamide
- Acetaminophen
Correct Answer: Cyclophosphamide
Q4. What is the primary cytological difference between a micronucleus caused by a clastogen versus an aneugen?
- Clastogen-induced micronuclei are always larger than aneugen-induced micronuclei
- Clastogen-induced micronuclei contain acentric chromosome fragments; aneugen-induced micronuclei contain whole chromosomes
- Aneugen-induced micronuclei stain completely differently from main nuclei
- Clastogen-induced micronuclei occur only in lymphocytes
Correct Answer: Clastogen-induced micronuclei contain acentric chromosome fragments; aneugen-induced micronuclei contain whole chromosomes
Q5. Which method helps distinguish whether micronuclei contain whole chromosomes or acentric fragments?
- Gram staining
- Centromere/kinetochore staining or FISH using centromeric probes
- ELISA for DNA fragments
- Western blot for tubulin
Correct Answer: Centromere/kinetochore staining or FISH using centromeric probes
Q6. During preparation of bone marrow metaphase spreads for chromosomal aberration analysis, what hypotonic solution is commonly used to swell cells?
- 0.9% NaCl
- 0.075 M KCl
- 100% ethanol
- PBS with EDTA
Correct Answer: 0.075 M KCl
Q7. Which feature is generally NOT scored as a chromosomal aberration in regulatory bone marrow CA tests according to common guidelines?
- Chromosome breaks
- Chromatid exchanges (e.g., dicentrics)
- Gaps (isolated achromatic lesions)
- Chromatid fragments
Correct Answer: Gaps (isolated achromatic lesions)
Q8. A marked decrease in the PCE/NCE ratio in a treated animal indicates which of the following?
- Increased erythropoiesis
- Bone marrow cytotoxicity or suppression
- Enhanced micronucleus induction specificity
- No effect on marrow
Correct Answer: Bone marrow cytotoxicity or suppression
Q9. Which reagent is commonly used to arrest dividing cells in metaphase for chromosomal aberration analysis?
- EDTA
- Colchicine or colcemid
- RNase A
- Trypsin
Correct Answer: Colchicine or colcemid
Q10. For a micronucleus assay in mice, a widely recommended minimum number of PCEs to score per animal for robust data is:
- 200 PCEs
- 2,000 PCEs
- 20,000 PCEs
- 50 PCEs
Correct Answer: 2,000 PCEs
Q11. Which staining method is routinely used for scoring micronuclei in bone marrow smears by bright-field microscopy?
- Giemsa or May-Grünwald-Giemsa staining
- Silver staining
- Gram staining
- Immunohistochemical DAB staining
Correct Answer: Giemsa or May-Grünwald-Giemsa staining
Q12. In in vivo chromosomal aberration tests, which tissue is most commonly used for scoring metaphases in rodents?
- Kidney
- Bone marrow
- Liver
- Skin
Correct Answer: Bone marrow
Q13. Which of the following is a hallmark criterion used to identify a micronucleus microscopically?
- Larger than the main nucleus
- Attached to the main nucleus by a thin filament
- Distinct from the main nucleus, round, with staining similar to main nucleus and usually less than one-third its diameter
- Only present in dead cells
Correct Answer: Distinct from the main nucleus, round, with staining similar to main nucleus and usually less than one-third its diameter
Q14. Which of the following agents is typically used as a positive control for inducing aneugenicity in genotoxicity assays?
- Mitomycin C
- Vincristine or vinblastine
- Urea
- Acetone
Correct Answer: Vincristine or vinblastine
Q15. Which analytical approach can increase throughput and reduce scorer bias in peripheral blood micronucleus assays compared with manual microscopy?
- Flow cytometry using fluorescence-labeled immature erythrocyte markers
- ELISA for micronuclear proteins
- Western blotting for histones
- Mass spectrometry
Correct Answer: Flow cytometry using fluorescence-labeled immature erythrocyte markers
Q16. Why is it important to include both negative (vehicle) and positive controls in an in vivo MN or CA study?
- Only to validate staining procedures
- To demonstrate assay sensitivity, verify test system responsiveness, and provide a basis for interpretation versus historical background
- Controls are optional for regulatory studies
- To increase the number of animals used without scientific reason
Correct Answer: To demonstrate assay sensitivity, verify test system responsiveness, and provide a basis for interpretation versus historical background
Q17. Which of the following best describes the origin of a chromatid-type aberration observed in metaphase spreads?
- Defect occurring after DNA replication affecting a single chromatid
- Numerical change of whole chromosomes only
- Artifact from slide staining only
- Occurs exclusively during anaphase
Correct Answer: Defect occurring after DNA replication affecting a single chromatid
Q18. According to typical regulatory practice, which of the following sampling times post-dose is commonly used to detect micronucleus induction in bone marrow of mice?
- 1 hour only
- 24 and/or 48 hours post-dose (often including 24–48 h and sometimes 72 h as appropriate)
- 2 weeks post-dose only
- 6 months post-dose
Correct Answer: 24 and/or 48 hours post-dose (often including 24–48 h and sometimes 72 h as appropriate)
Q19. Which limitation is most associated with the in vivo micronucleus test compared with detailed karyotyping for chromosomal damage?
- MN test provides detailed breakpoint information for structural rearrangements
- MN test cannot distinguish structural breakage from whole chromosome loss without additional centromere testing
- MN test identifies precise gene mutations
- MN test requires metaphase spreads
Correct Answer: MN test cannot distinguish structural breakage from whole chromosome loss without additional centromere testing
Q20. When preparing bone marrow slides for the chromosomal aberration assay, what fixative mixture is classically used to preserve chromosome morphology?
- Formalin only
- Methanol:acetic acid (3:1)
- Glutaraldehyde only
- Acetone:ether
Correct Answer: Methanol:acetic acid (3:1)
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