The Micronucleus Assay

In mammalian cells, small bodies of extra-chromatin denoted micronuclei can be observed in the cytoplasm at interphase and represent a rare abnormality consequent to genetic damage. Micronuclei are formed at the mitotic division by two different pathways (Fig. 1): (1) a pre-existing chromosome fragment lacking the centromere and, therefore, unable to move toward the spindle poles, can decondense into a micronucleus if excluded from the daughter nuclei (Fig. 1A); or (2) micronuclei can be derived from a chromosome lagging at anaphase, as described in Fig. 1B. Micronuclei are indirect but reliable indicators of chromosome damage: their frequency increases in either in vivo or in vitro cell populations exposed to different radiations or chemicals.

From: Methods in Molecular Biology, vol. 334: PRINS and In Situ PCR Protocols, Second Ed. Edited by: F. Pellestor © Humana Press Inc., Totowa, NJ

Fig. 1. The two pathways leading to micronucleus formation after a mitotic cell division. (A) top to bottom: a cell carrying a chromosome break (indicated by the black arrow) cannot control the segregation of the acentric fragment, which can be excluded from both daughter nuclei. In the cell progeny, depicted in the frame at bottom, one cell will carry a micronucleus. (B) top to bottom: a chromosome segregation error can occur during the mitotic cell division; if a chromatid is not able to keep or maintain contact with the spindle, it will lag at anaphase and possibly will be excluded from both daughter nuclei. In the cell progeny, depicted in the bottom frame, one cell will carry a micronucleus. Noteworthy, the two mechanisms lead to the same visible result, unless the micronucleus organization is not defined by molecular cytogenetics; for example, the micronucleus will contain the centromeric sequence only after pathway (B). For details see Fig. 2.

Fig. 1. The two pathways leading to micronucleus formation after a mitotic cell division. (A) top to bottom: a cell carrying a chromosome break (indicated by the black arrow) cannot control the segregation of the acentric fragment, which can be excluded from both daughter nuclei. In the cell progeny, depicted in the frame at bottom, one cell will carry a micronucleus. (B) top to bottom: a chromosome segregation error can occur during the mitotic cell division; if a chromatid is not able to keep or maintain contact with the spindle, it will lag at anaphase and possibly will be excluded from both daughter nuclei. In the cell progeny, depicted in the bottom frame, one cell will carry a micronucleus. Noteworthy, the two mechanisms lead to the same visible result, unless the micronucleus organization is not defined by molecular cytogenetics; for example, the micronucleus will contain the centromeric sequence only after pathway (B). For details see Fig. 2.

Thirty years ago, a micronucleus assay in in vivo proliferating rodent cells was proposed as a simple experimental procedure for the identification of chemical and physical agents affecting genome stability (1,2). In contrast to the direct chromosome aberration analysis in metaphase spreads, the micronucleus assay is fast and does not require skilled personnel. For these reasons, a great effort has been put to validate and improve the methodology with respect to the original proposal. At present, the micronucleus assay is a well-established and widely applied approach to evaluate the effects of potential mutagenic agents in several cell types in vivo (3) and in vitro (4).

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