The success of hematopoietic stem cells (HSCs) transplantation is limited due to the low number of HSCs received from donors. In vivo, HSCs reside within a specialized niche inside the 3D porous spongy bone. The natural environment in the niche is composed of structural proteins, glycosaminoglycans (GAGs) and soluble factors that control cells fate. However, the designed scaffolds for in vitro culture do not fairly recapitulate this microenvironment and cannot efficiently control HSCs fate. Here we report on the development of new omental ECM-based 3D macroporous sponges for hematopoietic cell culture. The scaffolds' structure, porosity and stability were characterized and optimized. Analysis of the biochemical content revealed that they were composed of collagens and GAGs, including sulfated GAGs. This morphology and composition enabled growth factors interaction with the sulfated GAGs, as indicated by the high loading capacity and release profile of three different hematopoietic niche factors. Finally, the ability of the ECM-based scaffolds to efficiently support the growth of hematopoietic cells by releasing stem cell factor (SCF) was demonstrated.