Sixteen new and seven recently published AMS radiocarbon dates from Guli Waabayo rock shelter in southern Somalia show repeated use of the site over a similar to 20,000-year period that spans most of Marine Isotope Stage (MIS) 2 similar to 29-14.5 thousand years ago (ka) and the MIS 1 African Humid Period similar to 14.5-6 ka. Improved methods of ostrich eggshell and tooth enamel sample preparation increase dating accuracy and provide the earliest radiometric evidence of Later Stone Age (LSA) occupation in the southeastern Horn of Africa. Bayesian analysis of 18 dates helps evaluate chronostratigraphic relationships at the site and define separate phases of site-use and formation. These data indicate two major periods of rock shelter occupation. The first spans MIS 2 and the distinctive LSA Eibian industry similar to 29-26 to similar to 15-11 cal ka. The second dates to the early/mid-Holocene and the LSA Bardaale industry similar to 11-9 to similar to 7.5-6 cal ka. This new Guli Waabayo chronology provides a framework for reconstructing and interpreting LSA hunter-gatherer adaptations to rapidly changing Late/terminal Pleistocene and early Holocene environments in what is now the semi-arid southeastern lowlands of the Horn of Africa.