Adult hematopoietic stem cells (HSCs) in the bone marrow (BM) are quiescent. Following perturbations, such as blood loss or infection, HSCs may undergo activation. Surprisingly, little is known about the earliest stages of HSCs activation. We utilize surface markers of HSCs activation, CD69 and CD317, revealing a response as early as 2 h after stimulation. The dynamic expression of HSCs activation markers varies between viral-like (poly-Inosinic-poly-Cytidylic) or bacterial-like (Lipopolysaccharide) immune stimuli. We further quantify dose response, revealing a low threshold, and similar sensitivity of HSCs and progenitors in the BM. Finally, we find a positive correlation between the expression of surface activation markers and early exit from quiescence. Our data show that the response of adult stem cells to immune stimulation is rapid and sensitive, rapidly leading HSCs out of quiescence.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13287-023-03377-6.