Optimizing chromosome yield: a comparative analysis of harvesting, preparation and waste recovery methods

In mitotic chromosome preparation, it is crucial to maximize chromosome yield for downstream cytogenetic analysis. Using HeLa cells as a model adherent cell, we assessed and compared the recovery of chromosomes from the entire process as well as the fraction of chromosomes that would generally become discarded in the standardly used trypsinization and mitotic-shake-off chromosome preparation methods. A higher chromosome yield for polyamine (PA) and methanol acetic acid (MAA) chromosomes was achieved using the mitotic-shake-off method compared to trypsinization. Moreover, mitotic arrest using colcemid or nocodazole gave similar PA and MAA chromosome yields in the commonly collected fractions. Interestingly, when comparing the fractions that would usually be discarded in the mitotic-shake-off, for colcemid-treated cells compared to nocodazole-treated cells, a greater number of PA chromosomes was recovered from the former. Our results show that chromosomes can be retrieved from the waste media. These recovered chromosomes display a suitable morphology in all chromosome preparations, suggesting that in conditions where high chromosome yields are required, utilizing the mitotic-shake-off method and recovering the generally discarded chromosome fraction together with the commonly used fraction would aid in maximizing chromosome yield.