Ablation characteristics of electrospun core-shell nanofiber by femtosecond laser.

This study examined the femtosecond laser ablation properties of core and shell polymers their relationship to the ablation characteristics of core-shell nanofibers. The single-pulse ablation threshold of bulk polycaprolactone (PCL) was measured to be 2.12J/cm(2) and that of bulk polydimethylsiloxane (PDMS) was 4.07J/cm(2). The incubation coefficients were measured to be 0.82±0.02 for PCL and 0.53±0.03 for PDMS. PDMS-PCL core-shell and pure PCL nanofibers were fabricated by electrospinning. The energy/volume of pure PCL and PDMS-PCL core-shell nanofiber ablation was investigated by measuring linear ablation grooves made at different scanning speeds. At large scanning speed, higher energy/volume was required for machining PDMS-PCL nanofiber than for PCL nanofiber. However, at small scanning speed, comparable energy/volume was measured for PDMS-PCL and PCL nanofiber ablation. Additionally, in linear scanned ablation of PDMS-PCL fibers at small laser pulse energy and large scanning speed, there were partially ablated fibers where the shell was ablated but the core remained. This was attributed to the lower ablation threshold of the shell material.

Design of a microchannel-nanochannel-microchannel array based nanoelectroporation system for precise gene transfection.

A micro/nano-fabrication process of a nanochannel electroporation (NEP) array and its application for precise delivery of plasmid for non-viral gene transfection is described. A dip-combing device is optimized to produce DNA nanowires across a microridge array patterned on the polydimethylsiloxane (PDMS) surface with a yield up to 95%. Molecular imprinting based on a low viscosity resin, 1,4-butanediol diacrylate (1,4-BDDA), adopted to convert the microridge-nanowire-microridge array into a microchannel-nanochannel-microchannel (MNM) array. Secondary machining by femtosecond laser ablation is applied to shorten one side of microchannels from 3000 to 50 µm to facilitate cell loading and unloading. The biochip is then sealed in a packaging case with reservoirs and microfluidic channels to enable cell and plasmid loading, and to protect the biochip from leakage and contamination. The package case can be opened for cell unloading after NEP to allow for the follow-up cell culture and analysis. These NEP cases can be placed in a spinning disc and up to ten discs can be piled together for spinning. The resulting centrifugal force can simultaneously manipulate hundreds or thousands of cells into microchannels of NEP arrays within 3 minutes. To demonstrate its application, a 13 kbp OSKM plasmid of induced pluripotent stem cell (iPSC) is injected into mouse embryonic fibroblasts cells (MEFCs). Fluorescence detection of transfected cells within the NEP biochips shows that the delivered dosage is high and much more uniform compared with similar gene transfection carried out by the conventional bulk electroporation (BEP) method.

Vascular wall engineering via femtosecond laser ablation: scaffolds with self-containing smooth muscle cell populations.

For tissue-engineered vascular grafts to reach their full potential, three-dimensional (3D) cellular micro-integration will be necessary. In this study, we utilize femtosecond laser ablation to produce microchannels inside electrospun polycaprolactone (PCL) scaffolds. These microchannels potentially provide spatially controlled cell distributions approaching those observed in vivo. The ability of such laser-ablated microchannels to direct cell seeding was evaluated. The dimensions chosen were 100 µm wide, 100 µm deep and 10 mm long. Femtosecond laser ablation successfully produced these microchannels in the scaffolds without substantially altering the ~900 nm diameter fibers. Flow within these microchannels was studied by injecting fluorescent polystyrene bead solutions. Direct measurement of bead motion yielded an inlet velocity of 2.78 cm s(-1). This was used for modeling two-dimensional (2D) flow using computational fluid dynamics to estimate flow profiles within the microchannel. Successful demonstrations of bead flow were followed by seeding of 500,000 human coronary artery smooth muscle cells (HCASMCs) in proliferative medium at a rate of ~500 µL min(-1). Confocal microscopy and scanning electron microscopy confirmed that the HCASMCs were seeded down the full 10-mm length of the microchannel and stayed within its boundaries. Both nuclei and F-actin were observed within the seeded cells. The presence of F-actin filaments shows that the cells were adhered strongly to the scaffold and remained viable throughout the culture. The concept of "vascular wall engineering" producing intricate cell seeding through microchannels produced via femtosecond laser ablation was validated.

Micropatterning and characterization of electrospun poly(ε-caprolactone)/gelatin nanofiber tissue scaffolds by femtosecond laser ablation for tissue engineering applications.

Experimental investigations aimed at assessing the effectiveness of femtosecond (FS) laser ablation for creating microscale features on electrospun poly(ε-caprolactone) (PCL)/gelatin nanofiber tissue scaffold capable of controlling cell distribution are described. Statistical comparisons of the fiber diameter and surface porosity on laser-machined and as-spun surface were made and results showed that laser ablation did not change the fiber surface morphology. The minimum feature size that could be created on electrospun nanofiber surfaces by direct-write ablation was measured over a range of laser pulse energies. The minimum feature size that could be created was limited only by the pore size of the scaffold surface. The chemical states of PCL/gelatin nanofiber surfaces were measured before and after FS laser machining by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy and X-ray photoelectron spectroscopy (XPS) and showed that laser machining produced no changes in the chemistry of the surface. In vitro, mouse embryonic stem cells (mES cells) were cultured on as-spun surfaces and in laser-machined microwells. Cell densities were found to be statistically indistinguishable after 1 and 2 days of growth. Additionally, confocal microscope imaging confirmed that spreading of mES cells cultured within laser-machined microwells was constrained by the cavity walls, the expected and desired function of these cavities. The geometric constraint caused statistically significant smaller density of cells in microwells after 3 days of growth. It was concluded that FS laser ablation is an effective process for microscale structuring of these electrospun nanofiber tissue scaffold surfaces.

Micropillar fabrication on bovine cortical bone by direct-write femtosecond laser ablation.

We investigated fabrication of cylindrical micropillars on bovine cortical bone using direct-write femtosecond laser ablation. The ablation threshold of the material was measured by single-pulse ablation tests, and the incubation coefficient was measured from linear scanned ablation tests. A motion system was programmed to apply multiple layers of concentric rings of pulses to machine pillars of various diameters and heights. The diameter of the top surface of the pillar was found to steadily decrease due to incubation of damage from successive layers of pulses during the machining process. Pillar top diameter was predicted based on a paraxial beam fluence approximation and single-pulse ablation threshold and incubation coefficient measurements. Pillar diameters predicted as successive layers of pulses were applied were well-matched to experiments, confirming that femtosecond laser ablation of the cortical bone was well-modeled by single-pulse ablation threshold measurements and an incubation coefficient.