Myeloma cells specifically localize in the bone marrow and rarely circulate in blood. To study whether this immobilization could be partially explained by the presence of constitutively activated integrins, particularly α4β1, we used the activation reporter HUTS-21 anti-β1 mAb. These analyses showed that β1 integrins on myeloma cells were moderately active and could be upregulated similarly to integrins on lymphoma or leukemia cells. Myeloma cells were also tested for their ability to attach to RGD-containing fibronectin fragments, a property of activated (but not resting) α4β1. Two cell lines adhered to these fragments and this was inhibited by anti-α5 but not by anti-α4 mAbs. These results show that myeloma cells bear low/moderately active α4β1 and support the notion that multiple interactions contribute to their localization in the bone marrow.