Biomimetic reconstruction of the hematopoietic stem cell niche for in vitro amplification of human hematopoietic stem cells.

Hematopoietic stem cell transplantation is successfully applied since the late 1950s; however, its efficacy still needs to be increased. A promising strategy is to transplant high numbers of pluripotent hematopoietic stem cells (HSCs). Therefore, an improved ex vivo culture system that supports proliferation and maintains HSC pluripotency would override possible limitations in cell numbers gained from donors. To model the natural HSC niche in vitro, we optimized the HSC medium composition with a panel of cytokines and valproic acid and used an artificial 3D bone marrow-like scaffold made of polydimethylsiloxane (PDMS). This 3D scaffold offered a suitable platform to amplify human HSCs in vitro and, simultaneously, to support their viability, multipotency and ability for self-renewal. Silicon oxide-covering of PDMS structures further improved amplification of CD34+ cells, although the conservation of naïve HSCs was better on non-covered 3D PDMS. Finally, we found that HSC cultivated on non-covered 3D PDMS generated most pluripotent colonies within colony forming unit assays. In conclusion, by combining biological and biotechnological approaches, we optimized in vitro HSCs culture conditions, resulting in improved amplification, multipotency maintenance and vitality of HSCs.

Single bead parallel synthesis and screening.

Microstructured silicon wafers were employed as miniaturized solid-phase reaction vessels as well as miniaturized micro titer plates. Employing piezoelectric drop-on-demand liquid jets, a combinatorial library of 256 Peptides was synthesized on single beads. The synthesis protocol was associated to the location in the silicon nano-well arrangement. Products were photolytically cleaved in the same well that was used for synthesis and subsequently interrogated for thrombin inhibition in a homogeneous competition assay. The assay procedure was based on drop-on-demand liquid delivery and laser induced fluorescence imaging. The novel format proved useful for the integration of both synthesis and screening into one platform, a prerequisite for an iterative, evolutionary approach towards drug discovery.

Microlenses as Amplification for CCD-Based Detection Devices for Screening Applications in Biology, Biochemistry, and Chemistry.

We describe the use of microlenses as amplification for CCD-based detection devices. The possible amplification of a signal in areaction chamber of a nanoplate is estimated with a first-order approximation. This value was proved with a commercially available microlens and with a specially constructed microlens array made of glass hemispheres. Possible applications of this approach to amplification are given.