Nearly 80% of patients with newly diagnosed bladder cancer present with superficial bladder tumors (confined to the bladder lining such as transitional cell carcinoma [90%], squamous cell carcinoma [6-8%], and adenocarcinoma[2%]) in stages Ta, Tis, or T1. Segmental cystectomy is one surgical treatment for patients who have a low-grade invasive tumor. Transposition of small intestine is a viable surgical treatment option. Success of the transplantation is also dependent upon removal of the entire SI mucosal layer. A Clark Spitfire Ti:Sapphire laser operating at 775 nm and 1 kHz repetition rate, was used to investigate the damage induced to fresh cadaveric porcine small intestinal mucosal epithelium. The laser was held constant at a focal spot diameter of 100 μm using a 200 mm focal point lens, with a power output maximum of 257 mW. A high resolution motorized X-Y-Z stage translated the SI tissue through the beam at 500 μm/sec with a line spacing of 50 μm. This produced a 50% overlap in the laser etching for each pass over a 1 cm x 1.5 cm grid. To determine if the mucosal lining of the SI was adequately removed, the targeted area was covered with 1% fluorescein solution for 30 seconds and then rinsed with phosphate buffered saline. Fluorescein staining was examined under UV illumination, to determine the initial degree of mucosal removal. Tissues were fixed and processed for light and scanning electron microscopy by standard protocols. Brightfield light microscopy of hematoxylin and eosin stained 4 μm thick cross sections, scanning electron microscopy were examined to determine the degree of mucosal tissue removal. Clear delineation of the submucosal layer by fluorescein staining was also observed. The Ti:Sapphire laser demonstrated precise, efficient removal of the mucosal epithelium with minimal submucosal damage.
Laser micromachining combined with digital printing allows rapid prototyping of complex bioreactors with reduced fabrication times compared to multi-mask photolithography. Microfluidic bioreactors with integrated optical waveguides for diagnostics have been fabricated via ultrashort pulse laser micromachining and digital printing. The microfluidic channels are directly laser machined into poly(dimethylsiloxane) (PDMS) silicone elastomer. Multimode optical waveguides are formed by coating the PDMS with alternating refractive index polymer layers and laser machining to define the waveguide geometry. Tapered alignment grooves are also laser machined to aid in coupling optical fibers to the waveguides. Three-dimensional (3-D) bio-scaffold matrices comprising liquid solutions that can be selectively and rapidly gelled are digitally printed inside the bioreactors and filled with nutrient rich media and cells. This paper will describe the maskless fabrication of complex 3-D bioreactors and discuss their performance characteristics.
Novel devices can be relatively simple in theory and modeling, but difficult and many times unfeasible to fabricate in a traditional cleanroom environment. We have developed a CAD/CAM tool capable of integrating multiple materials in the electronic, photonic, and biological regimes for applications in both MEMS and BioMEMS devices. Some materials are known and more fully characterized, such as thick film resistors or conductors, while other materials such as biodegradable scaffolding are new but showing promise to realize heterogenous tissue engineered constructs and drug delivery devices. The tool does not discriminate, but rather places these materials in specified locations with precision volumetric control, gently, conformally, and in 3-D. This paper will describe the enabling aspect of true 3-D maskless fabrication as well as describe multiple device structures and demonstrations.
A Ti:Sapphire laser operating at 800 nm and 1 kHz repetition rate, was used to investigate the damage induced to fresh cadaveric porcine tissues. The laser was held constant at a focal spot diameter of 100 μm for pulse widths varying from 120-femtoseconds to 7-nanoseconds yielding a maximum fluence of 12.7 J/cm2 irradiation. Polarization optics were used to reduce the energy per pulse to well below tissue ablation threshold fluences. Hollow silica waveguides with a silver inner coating and bore diameters of 300, 500, 750 and 1000 μm were also used for the Ti:Sapphire laser with output pulses <150 fs duration and energy up to 700 μjoules. A high resolution motorized X-Y-Z stage translated the tissue through the beam at 1 mm/sec. A Luxar Novapulse CO2 surgical laser was used as a standard for comparison. Tissues were processed for light, scanning and transmission electron microscopy by standard protocols. Tissue samples were examined for tissue removal rates, thermal damage to adjacent tissue, and cellular disruption for equivalent fluence levels. The Ti:Sapphire laser demonstrated an increase in removal rate along with a decrease in thermal damage as the pulse widths approached the femtosecond regime for a constant fluence. With femtosecond pulses, ablation still occurred below fluences of 2 J/cm2. However, for nanosecond pulses, ablation no longer occurred, showing a decrease in ablation threshold as the pulse width decreases. Because of the reduced thermal effects compared to nanosecond pulses, ultrafast lasers may offer a solution to more precise tissue removal with less damage to surrounding cells as compared to more conventional surgical laser systems.
Using a Ti:Sapphire laser operating at 800nm and a repetition rate of 1 kHz, we investigated the damage induced to fresh cadaveric porcine liver after laser irradiation for pulse widths of 120-fs, 8ps, and 7-ns. The laser was held constant at a focal spot diameter of 100μm yielding a maximum fluence of 9J/cm2. Then, using polarization optics, the energy per pulse was controlled to well below ablation threshold fluences. The tissue samples were translated under the laser via 0.1μm resolution encoded X-Y-Z motorized stages. After irradiation and fixation, we evaluated the tissues using brightfield light microscopy of Hematoxylin and Eosin stained 4 μm thick cross sections, scanning electron microscopy, and transmission electron microscopy. The tissue samples were examined for both removal rates of material, thermal damage to surrounding tissue, and cell disruption for equivalent fluence levels across the temporal range. We found an increase in removal rate along with a decrease in thermal damage as the pulse widths approached the femtosecond regime for a constant fluence. With femtosecond pulses, ablation still occurred below fluences of 2J/cm2. However, for nanosecond pulses, ablation no longer occurred, showing a decrease in ablation threshold as the pulse width decreases. Because of the reduced thermal effects compared to nanosecond pulses, ultrafast lasers may offer a solution to more precise tissue removal with less damage to surrounding cells.
Tissue ablation with pulses in the femtosecond regime is generally more efficient and causes less collateral and thermal damage to the surrounding tissue compared to ablation with longer pulsewidths. A compact, flexible fiber delivery system that could transmit these pulses would be advantageous over free-space beam delivery, since it would allow ultrashort pulse tissue ablation in vivo. However, the extremely high intensities associated with ultrashort pulses have deleterious effects in conventional silica fibers such as nonlinearities and fiber damage. Hollow silica waveguides with a silver inner coating essentially guide the pulses in air, thereby avoiding many of these problems. The transmission characteristics of four hollow waveguides with bore diameters of 300, 500, 750 and 1000μm and lengths up to 1m were tested using pulses from a femtosecond regime Ti:Sapphire laser operating with input pulses <150fs duration and energy up to 700 microjoules at a repetition rate of 1kHz. Coupling was primarily to the HE11 mode and straight and bending losses were measured. Beam profiles were also taken at the output of straight and bent waveguides. Autocorrelation measurements show minimal pulse broadening for straight waveguides and increasing pulsewidth with waveguide bend. Diffractive micro-optics were used to focus the output and ablation of fresh cadaveric porcine liver and heart tissues was accomplished using an x-y-z translational stage moving at 1 mm/second. Targeted tissues were then processed for light and electron microscopic examination. Light and scanning electron microscopy demonstrated near a-thermal ablation with depth correlating to energy application.
Multimode optical waveguides have been fabricated by dispensing photopolymers onto various substrates using direct-write technologies. Fine-tuning is achieved by micromachining the waveguides using a femtosecond-regime pulsed laser. Propagation losses are determined using the cutback method. A 2 × 2 coupler and a 1 × 8 splitter are demonstrated.