Introduction: High concentrations of hexabromocyclododecane (HBCD) sometimes recorded in free-range hens' eggs are thought to be due to soil ingestion. Of the three stereoisomers of HBCD (α-, β-, and γ-HBCD), γ-HBCD is the main component in the commercial mixture, as well as in environmental matrices, whereas the isomer profile is α-dominated in biota. In fish and in mammals, this shift is thought to be due to a rapid elimination of γ-HBCD and to its bioisomerization to the more persistent α-HBCD. The aim of the current controlled study was to better understand the fate of ingested HBCD in laying hens. The isomer profile in soil being γ-dominated, excretion kinetics of γ-HBCD into egg yolk, and accumulation in liver and in abdominal fat were investigated. Materials and methods: Forty-eight laying hens were individually housed and fed with a spiked diet containing 1.1-ng γ-HBCD per gram for 21 days and with a clean diet for the following 18 days. Hens were sequentially slaughtered throughout the 39-day experiment. α-, β-, and γ-HBCD were analyzed in egg yolk, in abdominal fat, and in liver by LC-MS/MS. α- and γ-HBCD were quantified in the three tissues, while β-HBCD was never quantified. Results and conclusion: Kinetics of the two isomers suggests that γ-HBCD is rapidly biotransformed and eliminated, and partly isomerized into the more persistent α-HBCD. Carry-over rate of ingested γ-HBCD to egg yolk was estimated at 1.2%. Estimated half-lives of γ-HBCD in egg yolk, in abdominal fat, and in liver were 2.9, 13, and 0.41 days, respectively.