We studied sex allocation of isogenic unicellular homothallic centric diatom with the single-cell resolution under tightly controlled induction conditions. We designed a microfluidic system that allows to track cell divisions and sexual differentiation of multiple lineages in parallel by trapping cells in an array of micrometre-size chambers. Our study revealed that sex allocation of individual mother cells is affected by three factors: cell lineage, cell size and cell density. First, the sibling cell pairs showed the bias toward the same choice of sexual differentiation fates. Second, we found a significant negative correlation between the size of mother cell area and a gametangium sex ratio of 2 sibling cells. Third, the male-biased sex ratio declined as cell density within the microchamber increases. Together, these results have characterized multiple factors affecting the sex allocation of a homothallic unicelluar organism.