Brain extracellular matrix (ECM) is often overlooked in vitro brain tissue models, despite its instructive roles during development. Using developmental stage-sourced brain ECM in reproducible 3D bioengineered culture systems, we demonstrate enhanced functional differentiation of human induced neural stem cells (hiNSCs) into healthy neurons and astrocytes. Particularly, fetal brain tissue-derived ECM supported long-term maintenance of differentiated neurons, demonstrated by morphology, gene expression and secretome profiling. Astrocytes were evident within the second month of differentiation, and reactive astrogliosis was inhibited in brain ECM-enriched cultures when compared to unsupplemented cultures. Functional maturation of the differentiated hiNSCs within fetal ECM-enriched cultures was confirmed by calcium signaling and spectral/cluster analysis. Additionally, the study identified native biochemical cues in decellularized ECM with notable comparisons between fetal and adult brain-derived ECMs. The development of novel brain-specific biomaterials for generating mature in vitro brain models provides an important path forward for interrogation of neuron-glia interactions.

15Brain extracellular matrix (ECM) is often overlooked in vitro brain tissue models, despite its 16 instructive roles during development. Using developmental stage-sourced brain ECM in reproducible 3D 17 bioengineered culture systems, we demonstrate enhanced functional differentiation of human induced 18 neural stem cells (hiNSCs) into healthy neurons and astrocytes. Particularly, fetal brain tissue-derived 19 ECM supported long-term maintenance of differentiated neurons, demonstrated by morphology, gene 20 expression and secretome profiling. Astrocytes were evident within the second month of differentiation, 21 and reactive astrogliosis was inhibited in brain ECM-enriched cultures when compared to unsupplemented 22cultures. Functional maturation of the differentiated hiNSCs within fetal ECM-enriched cultures was 23 confirmed by calcium signaling and unsupervised cluster analysis. Additionally, the study identified 24 native biochemical cues in decellularized ECM with notable comparisons between fetal and adult brain-25 derived ECMs. The development of novel brain-specific biomaterials for generating mature in vitro brain 26 models provides an important path forward for interrogation of neuron-glia interactions. 27

Breakthrough cell therapies for the treatment of cancers require the separation of specific cells, such as T cells, from the patient's blood. Current cell therapy processes rely on magnetic separation, which adds clinical risk and requires elevated manufacturing controls due to the added foreign material that constitutes the magnetic beads. Acoustophoresis, a method that uses ultrasound for cell separation, has demonstrated label-free enrichment of T cells from blood, but residual other lymphocytes limit the ultimate purity of the output T cell product. Here, to increase the specificity of acoustophoresis, we use affinity reagents to conjugate red blood cells with undesired white blood cells, resulting in a cell–cell complex (rosette) of increased acoustic mobility. We achieve up to 99% purity of T cells from blood products, comparable to current standards of magnetic separation, yet without the addition of separation particles.

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