Background: Natural Killer (NK) cells play a critical role in host defense. Studying human NK immunobiology is mainly focused on using in vitro assays with limited NK cells from peripheral blood. It is challenging to study human NK cell biology in vivo due to potential ethical issues in human study and the lack of suitable animal models. Developing a suitable animal model to study human NK cell biology in vivo is critical to support NK-based clinical immunotherapy. Results: Here, we develop a novel method to study human NK cells in vivo by using hu-BLT (humanized bone marrow-liver-thymus) mice that constitutively express human IL-15 (henceforth, hu-BLT-IL15). We also compare human NK cells between hu-BLT-IL15 and hu-BLT mice without IL-15 expression by a newly developed approach for the rapid propagating of primary human NK cells from various sources (including peripheral blood, spleen, and bone marrow). NK cells from hu-BLT-IL15 show superior number, purity, and cytotoxicity (including natural cytotoxicity and antibody-dependent cellular cytotoxicity [ADCC]), compared with NK cells from hu-BLT. Unexpectedly, we also identify a significantly increased percentage of NK-like T cells (CD3+ CD16+ CD56+) from hu-BLT-IL15, indicating that IL-15 signaling enhances both NK and NKT cell development. Conclusions: A better understanding of the immunobiology of the NK-like T cells in the hu-BLT-IL15 mouse model may provide critical information for determining the clinical value of these cells in predicting disease progression. Thus, we propose that the hu-BLT-IL15 mouse model in combination with the 721.221-mIL21 feeder cell expansion system can serve as a superior model to study human NK and NK-like T cells in comparison with hu-BLT.