Digital holographic microscopy (DHM) has a wide variety of applications in biomedical sciences. Here we examine the use of an off-axis holographic system to monitor bacterial growth in commercially available ibidi micro-fluidic chambers. The imaged volume spans roughly 120 x 120 x 200 mu m(3), at 50x magnification. By use of an off-axis reference wave and some standard numerical filtering operations it is possible to recover the complex amplitude and thus the phase of the object wave. Two strains of bacteria are used in the research study: Escherichia coli and Staphylococcus warneri with dimensions around 1-4 microns. Examining these samples, showed that through the use of phase compensation techniques, the method can sustain an average Signal-to-Noise ratio (SNR) of around 5, allowing for the use of a simple thresholding algorithm for cell counting. In addition, the cell count dynamics determined using this method reflect the exponential increase of dilution steps very well, when compared against hypothetical cell counts. Details of the system operation, including a discussion of the signal processing routines, together with modifications to the general optic design of DHM systems are presented.

A rapid, precise, and viability-retaining method for cytoplasmic molecule delivery is highly desired for cell engineering. Routine methods suffer from low throughput, lack of selectivity, requirement of helper compounds, predominant endosomal delivery, and/or are restricted to specific molecule classes. Photonic cell manipulation bears the potential to overcome these drawbacks. Here we investigated mammalian cell manipulation by single sub-nanosecond laser pulses. Axial beam waist positioning close to a cell monolayer induced culture vessel damage and zones of cell ablation. Cells at margins of ablation zones exhibited uptake of membrane-impermeant fluorophores and GFP expression plasmids. Increasing Rayleigh-length and beam waist diameter reduced the sensitivity to axial defocusing and resulted in robust molecule transfer. Serial application of single pulses focused over a moving cell monolayer yielded quantitative molecule transfer to cells at rates up to 40%. Our results could be basic to spatially and temporally controlled single laser pulse-mediated marker-free high throughput cell manipulation.

A novel diode pumped Tm:YAG laser (Pantec Biosolutions AG) with a more flexible temporal pulse regime is available. This study includes first experiments with model stones on the influence of the pulse regime on the fragmentation rate. For this purpose, ablation experiments were performed on rectangular model stones (BEGO, 40 mm x 10 mm x 5 mm). The laser beam was coupled into a 270 μm light guide and the distal fiber end was positioned in <50 μm to the stone surface. This was located in a basin filled with water, which was moved horizontally with the stone at constant velocity by a computer-controlled translation stage. The ablation rate and the ablation efficiency were determined by subsequent measurement of the depth and width of the resulting crater. The experiments were performed with varying parameters of the novel pulse regime and, for comparison, with standard laser settings. The experiments show significant differences in bubble dynamics and shape, depending on the temporal pulse regime. Lowest values for propulsion were measured for the standard pulse regime. The measured and calculated values for ablation depth, -rate and -efficiency are comparable for all investigated pulse regimes. Especially the ablation efficiency is quite high compared to values which were calculated from published data. In conclusion, these preliminary results show a high potential of the diode pumped Tm:YAG laser with novel laser driver for variable and high efficient lithotripsy. The wide range of available peak power should allow fragmentation as well dusting with the same laser system.

Objectives: A sufficient histological evaluation is a key pillar in oncological treatment, especially in situations of cancer of unknown primary. CO2 laser technology is used in clinical routine of soft tissue surgery because of its cutting quality and availability. Diode pumped solid state Er(bium):YAG laser systems promise a higher cutting efficiency and minor thermal damages. The aim of this study was to compare both laser systems with respect to their suitability for cutting soft tissue. Methods: A setup was realized which enables comparable experiments with the clinical CO2 laser (AcuPulse 40ST DUO, Lumenis) and the Er:YAG laser system (DPM 40, Pantec Biosolutions AG). Fresh mucosal samples of porcine tongues were used to determine the influence of laser power and sample velocity on cutting depth and thermal damage width for both lasers. In addition, for the Er:YAG laser, the influence of the pulse repetition rate was examined additionally. For analysis, images of histological sections were taken. Results: In all experiments, the Er:YAG laser shows a significantly higher cutting depth (P < 0.0001) and less thermal damage width (P < 0.0001) than the CO2 laser. For example, at an average power of 7.7 W and a sample velocity of 5 mm/s the Er:YAG laser shows a mean cutting depth of 1.1 mm compared to the CO2 laser with 500 μm. While the Er:YAG laser shows a mean thermal damage width of 70 μm compared to 120 μm. Furthermore, the Er:YAG enables the adjustment of the cutting depth and thermal damage width by varying the irradiation parameters. A decrease of the repetition rate leads to a reduction of thermal damage. For example, a repetition rate of 100 Hz results in a thermal damage width of 46 μm compared to 87 μm at 800 Hz at an average power of 7.7 W and a cutting velocity = 5 mm/s while a homogenous cutting quality can be achieved. Conclusions: In conclusion, the results of these ex vivo experiments demonstrate significant advantages of the diode pumped Er:YAG laser system for soft tissue ablation compared to the CO2 laser, in particular regarding cutting efficiency and thermal damage width.

For some time now, the diode-pumped Er:YAG laser (Pantec Biosolutions AG) has been available. Due to its high repetition rate of up to 2 kHz, it shows an excellent cutting quality and efficiency in soft tissue. Now, due to the novel special pulse regime, significantly longer effective pulse duration and higher pulse energy of up to 1.5 J is available. The aim of the present study was to evaluate the ablation quality, efficiency and thermal side effects of the new pulse regime. For this purpose, in-vitro experiments were performed on slices of teeth and pig bones with varying laser irradiation parameters. The laser beam was coupled into a sapphire fiber and the fiber end was imaged onto an 800 µm spot on the sample. The sample was shifted with a computer-aided movement unit at different speeds during irradiation. A water spray with 6 ml/min was used for moistening. After irradiation, the resulted ablation quality was recorded under the light microscope and the ablation depth and width were measured. The ablation efficiency in super pulse mode is comparable on enamel and exceeds the values in dentin for standard pulse mode. The maximum ablation efficiency on bone is 0.20 mm3/J and is on average approx. 18,2% above the values achieved in standard operation. The ablation quality and the thermal injury achieved with the novel pulse regime is comparable to the results obtained with the standard operation of the diode-pumped Er:YAG laser. Overall, the tests show that the diode-pumped Er:YAG laser with the novel pulse regime and higher effective pulse energy allows larger spot size and with this more homogeneous ablation.

This paper presents a universal point-of-care system for fully automated quantification of human T-cell lymphotropic virus type 1 (HTLV-1) proviral load, including genomic RNA, based on digital reverse RNA transcription and c-DNA amplification by MD LAMP (mediator displacement loop-mediated isothermal amplification). A disposable microfluidic LabDisk with pre-stored reagents performs automated nucleic acid extraction, reaction setup, emulsification, reverse transcription, digital DNA amplification, and quantitative fluorogenic endpoint detection with universal reporter molecules. Automated nucleic acid extraction from a suspension of HTLV-1-infected CD4+ T-lymphocytes (MT-2 cells) yielded 8 ± 7 viral nucleic acid copies per MT-2 cell, very similar to the manual reference extraction (7 ± 2 nucleic acid copies). Fully automated sample processing from whole blood spiked with MT-2 cells showed a comparable result of 7 ± 3 copies per MT-2 cell after a run time of two hours and 10 min.

This work describes the development of a micro-integrated multi-spectral imaging system for hyperspectral imaging (HSI). The aim is to employ polymeric optical components to increase integration density and reduce weight while providing improved functionality compared to classical HSI camera systems. Economic advantages of both, simplified assembly by direct integration of mounting features and the ability of mass production of polymer optics are being addressed. An integration strategy into a camera system is developed, providing the required features for micro-assembly of all components. Methods for micro assembly of a polymeric microlens array (MLA) with 12.000 micro lenses in relation to the positioning of a Charge-Coupled Device (CCD) chip as well as a diffraction grating are applied. The direct integration of mounting features into the MLA allows for ultra-compact assembly with a reduced number of assembly processes. Furthermore, it enables the alignment of spectral lines to the CCD pixels, thus optimizing the number of spectra mapped on a single chip. The functionality of system is demonstrated, presenting a technological improvement for a large variety of applications, which may profit from the availability of such a system in terms of simplicity, costs, size and weight.

We present an automated point-of-care testing (POCT) system for rapid detection of species- and resistance markers in methicillin-resistant Staphylococcus aureus (MRSA) at the level of single cells, directly from nasal swab samples. Our novel system allows clear differentiation between MRSA, methicillin-sensitive S. aureus (MSSA) and methicillin-resistant coagulase-negative staphylococci (MR-CoNS), which is not the case for currently used real-time quantitative PCR based systems. On top, the novel approach outcompetes the culture-based methods in terms of its short time-to-result (1 h vs. up to 60 h) and reduces manual labor. The walk-away test is fully automated on the centrifugal microfluidic LabDisk platform. The LabDisk cartridge comprises the unit operations swab-uptake, reagent pre-storage, distribution of the sample into 20 000 droplets, specific enzymatic lysis of Staphylococcus spp. and recombinase polymerase amplification (RPA) of species (vicK) - and resistance (mecA) -markers. LabDisk actuation, incubation and multi-channel fluorescence detection is demonstrated with a clinical isolate and spiked nasal swab samples down to a limit of detection (LOD) of 3 ± 0.3 CFU μl-1 for MRSA. The novel approach of the digital single cell detection is suggested to improve hospital admission screening, timely decision making, and goal-oriented antibiotic therapy. The implementation of a higher degree of multiplexing is required to translate the results into clinical practice.

Flashlamp pumped Erbium lasers are successfully used clinical for soft and hard tissue ablation. Especially for soft tissue ablation the limited repetition rate is a disadvantage (bleeding; perforation instead of cutting). Now diode pumped solid state (DPSS) Er:YAG laser systems (Pantec Engineering AG) are available, with mean laser power up to 50 W and pulse repetition rate up to 2 kHz. The aim of this study is to investigate the potential of this laser system for increased and defined soft tissue coagulation/manipulation at various irradiation parameters, in particular at repetition rates exceeding 100 Hz. Firstly, an appropriate experimental set-up was realized with laser system, focusing unit, computer-controlled linear stage with sample holder and shutter unit to move the sample (fresh chicken breast) with a defined velocity while irradiation by various laser parameters. While irradiation the tissue effects were recorded by a video camera, adapted on a surgical microscope. After irradiation, the samples were analyzed by light microscopy. In addition, histological sections were prepared and microscopically analyzed. Mainly depending on the fluence, the thermal effect can be limited to coagulation without carbonization. In addition, tissue melting can be observed. The coagulation depth increases with increasing pulse repetition rate and decreasing movement velocity from about 30 µm to above 1 mm. In conclusion, the results of the in vitro studies show that the diode pumped, pulsed Er:YAG laser has the potential to provide one system both, for high efficient hard and soft tissue ablation as well as for soft tissue coagulation and fusion.

Spatially and temporally controlled drug delivery is important for implant and tissue engineering applications, as the efficacy and bioavailability of the drug can be enhanced, and can also allow for drugging stem cells at different stages of development. Long-term drug delivery over weeks to months is however difficult to achieve, and coating of 3D surfaces or creating patterned surfaces is a challenge using coating techniques like spin- and dip-coating. In this study, mesoporous films consisting of SBA-15 particles grown onto silicon wafers using wet processing were evaluated as a scaffold for drug delivery. Films with various particle sizes (100 – 900 nm) and hence thicknesses were grown onto OTS-functionalized silicon wafers using a direct growth method. Precise patterning of the areas for film growth could be obtained by local removal of the OTS functionalization through laser ablation. The films were incubated with the model drug DiO, and murine myoblast cells (C2C12 cells) were seeded onto films with different particle sizes. Confocal laser scanning microscopy (CLSM) was used to study the cell growth, and a vinculin-mediated adherence of C2C12 cells on all films was verified. The successful loading of DiO into the films was confirmed by UV-vis and CLSM. It was observed that the drugs did not desorb from the particles during 24 hours in cell culture. During adherent growth on the films for 4 h, small amounts of DiO and separate particles were observed inside single cells. After 24 h, a larger number of particles and a strong DiO signal were recorded in the cells, indicating a particle mediated drug uptake. A substantial amount of DiO loaded particles were however attached on the substrate after 24 making the films attractive as a long-term reservoir for drugs on e.g. medical implants.

Er:YAG lasers (3 mu m) allow efficient bone ablation caused by the strong absorption in water. Unfortunately, there are only a few and comparable expensive fiber materials for this wavelength available which are suitable for high laser power. The bone ablation efficiency of the Tm:YAG laser is minor (2 mu m) but inexpensive silica fibers can be used. The aim of this study was to investigate the bone ablation, using novel diode pumped high power Er:YAG (laser power 40W) and Tm:YAG laser system (60W) and adaptive fiber delivery systems. Expected advantage of these lasers is the longer lifetime of the fibers because of the high repetition rate and low pulse energy compared to the flash lamp pumped laser systems. The bare fiber output ends of a sapphire fiber (Er:YAG laser) and of a silica fiber (Tm:YAG laser) were attached under water and a water filled container including the fixed sample (bovine bone slices) was moved by a computer controlled translation stage. In a second set- up we provided a focusing unit and appropriate water spray unit. The generated cut kerfs were analyzed by light microcopy and laser scanning microscopy. The results show that with the diode pumped Er:YAG laser and sapphire fiber a particular high efficient bone ablation (>0.16mm(2)/J) is possible both with bare fiber under water and focusing unit with water spray. The higher power of the Tm:YAG laser also results in high ablation rates but causes enlarged thermal damages. In conclusion, this study demonstrates that efficient bone ablation is possible with both diode pumped laser systems. In terms of efficiency the Er:YAG laser is outstanding. The Tm:YAG laser also allows fast bone ablation, provided that the thermal impact is limited by effective cooling and high movement velocity of the laser spot, for example by using an automatic scanner.

Several studies have shown the potential of the diode pumped Er: YAG laser for medical applications. Benefits are the efficient and precise ablation of both hard and soft tissue with adjustable thermal effects.Aim of this study is the investigation of the ablation process on dentin and enamel using the diode pumped Er: YAG laser even at higher mean laser power and two different handpieces (free beam vs. contact).At first the laser radiation of the diode pumped Er: YAG laser (DPM-25, PANTEC Engineering AG) was coupled into a standard dental fiber delivery system (KEY3 laser, KaVo GmbH) with the two handpieces for cavity preparation. An appropriate experimental setup was realized including a computer controlled translation stage with sample holder to move the dentin or enamel slides of extracted human teeth with a defined velocity while irradiation by various laser parameters. After irradiation, the cut kerfs were analyzed by light microscopy and laser scanning microscopy.The results show a good ablation quality for both handpieces and a higher ablation rate in dentin and enamel using the non-contact handpiece compared to the contact handpiece. The calculated ablation efficiency is remarkable high (dentin: 0.094-0.172 mm 3/J, enamel: 0.025-0.093 mm 3/J) and exceeds all published values. The results of the high-speed camera observations explain the differences between the handpieces.In conclusion these experiments with the diode pumped Er: YAG laser system demonstrate its ability for efficient and very fast cavity preparation.

Flashlamp pumped Er:YAG lasers are successfully used clinically for both precise soft and hard tissue ablation. Since several years a novel diode pumped Er:YAG laser system (Pantec Engineering AG) is available, with mean laser power up to 40 W and pulse repetition rate up to 1 kHz. The aim of the study was to investigate the suitability of the laser system specifically for stapedotomy. Firstly an experimental setup was realized with a beam focusing unit and a computer controlled translation stage to move the samples (slices of porcine bone) with a defined velocity while irradiation with various laser parameters. A microphone was positioned in a defined distance to the ablation point and the resulting acoustic signal of the ablation process was recorded. For comparison, measurements were also performed with a flash lamp pumped Er:YAG laser system. After irradiation the resulting ablation quality and efficacy were determined using light microscopy. Using a high speed camera and “Töpler-Schlierentechnik” the cavitation bubble in water after perforation of a bone slice was investigated. The results show efficient bone ablation using the diode pumped Er:YAG laser system. Also a decrease of the sound level and of the cavitation bubble volume was observed with decreasing pulse duration. Higher repetition rates lead to a slightly increase of thermal side effects but have no influence on the ablation efficiency. In conclusion, these first experiments demonstrate the high potential of the diode pumped Er:YAG laser system for use in middle ear surgery.

The aim of this study was to determine the potential of a novel diode-pumped Er:YAG laser for phacoemulsification in basic experimental investigations. An appropriate experimental setup was created, including a translation stage for sample movement, a sample holder, a water spray for sample humidification and a surgical microscope with a CCD camera for video documentation. The analysis of the laser cuts and histological sections was done by light microscopy. As samples porcine eye lenses hardened by formalin were used. In ablation experiments with different spot diameters and radiant powers and a constant repetition rate νr = 200 Hz the maximum ablation depths of (4.346 ± 0.044) mm have reached at (Ø = 480 μm, Φ = 24.15 W) with a maximum extend of thermal damage of (0.165 ± 0.030) mm. The average ablation efficiency is 0.241 mm3/J. With a spot diameter of 308 μm the maximum ablation depth is (4.238 ± 0.040) mm at 24.65 W with a mean ablation efficiency of 0.293 mm3/J. The extend of the thermally damaged region is (0.171 ± 0.024) mm at this laser power. Using a sapphire cylinder with a diameter of 412 μm (length 38.5 mm) in direct tissue contact with water spray for sample humidification the ablation depth reaches (1.017 ± 0.074) mm at 4.93 W and (1.840 ± 0.092) mm at 9.87 W with a mean efficiency of 0.261 mm3/J. A thermal damage zone of (0.064 ±0.024) mm at 9.87 W was measured. Additionally, at this high power, a progressive contamination and destruction of the cylinder end facet was observed. In conclusion, the investigations show that the diode-pumped Er:YAG laser has considerable potential for cataract surgery.

Flash lamp pumped Er: YAG-lasers are used in clinical practice for dental applications successfully. As an alternative, several diode pumped Er: YAG laser systems (Pantec Engineering AG) become available, with mean laser power of 2W, 15W, and 30W.The aim of the presented study is to investigate the potential of the 2W Er: YAG laser system for oral surgery.At first an appropriate experimental set-up was realized with a beam delivery and both, a focusing unit for non-contact tissue cutting and a fiber tip for tissue cutting in contact mode. In order to produce reproducible cuts, the samples (porcine gingiva) were moved by a computer controlled translation stage. On the fresh samples cutting depth and quality were determined by light microscopy. Afterwards histological sections were prepared and microscopically analyzed regarding cutting depth and thermal damage zone.The experiments show that low laser power <= 2W is sufficient to perform efficient oral soft tissue cutting with cut depth up to 2 mm (sample movement 2mm/s). The width of the thermal damage zone can be controlled by the irradiation parameters within a range of about 50 mu m to 110 mu m. In general, thermal injury is more pronounced using fiber tips in contact mode compared to the focused laser beam.In conclusion the results reveal that even the low power diode pumped Er: YAG laser is an appropriate tool for oral surgery.

It is well known that flashlamp pumped Er:YAG lasers allow efficient bone ablation due to strong absorption at 3 mu m by water. Preliminary experiments revealed also a newly developed diode pumped Er: YAG laser system (Pantec Engineering AG) to be an efficient tool for use for bone surgery.The aim of the present in vitro study is the investigation of a new power increased version of the laser system with higher pulse energy and optimization of the treatment set-up to get high cutting quality, efficiency, and ablation depth.Optical simulations were performed to achieve various focus diameters and homogeneous beam profile. An appropriate experimental set-up with two different focusing units, a computer controlled linear stage with sample holder, and a shutter unit was realized. By this we are able to move the sample (slices of pig bone) with a defined velocity during the irradiation. Cutting was performed under appropriate water spray by moving the sample back and forth. After each path the ablation depth was measured and the focal plane was tracked to the actual bottom of the groove. Finally, the cuts were analyzed by light microcopy regarding the ablation quality and geometry, and thermal effects.In summary, the results show that with carefully adapted irradiation parameters narrow and deep cuts (ablation depth > 6mm, aspect ratio approx. 20) are possible without carbonization.In conclusion, these in vitro investigations demonstrate that high efficient bone cutting is possible with the diode pumped Er: YAG laser system using appropriate treatment set-up and parameters.

The successful clinical application of the Er:YAG-laser in dentistry is well known, documented by numerous published studies. These lasers are flash lamp pumped systems and emit pulses of typically some 100 mu s duration with energies of up to 1 J. Pulse repetition rates can reach up to 100Hz, and mean powers are up to about 8W. As an alternative to these laser systems recently a novel diode pumped Er:YAG laser system (Pantec Engineering AG) became available. This laser can provide a pulse repetition rate up to 2kHz and a mean laser power up to 15W. The aim of the presented study is to investigate the effect of this laser system on dental hard and soft tissue at various irradiation parameters, particular at repetition rates more than 100 Hz.At first an appropriate experimental set-up was realized with a beam delivery and focusing unit, a computer controlled stepper unit with sample holder, and a shutter unit. The stepper unit allows to move the samples ( dentin or enamel slides of extracted human teeth, chicken breast, pig bone) with a defined velocity during irradiation by various laser parameters. For rinsing the sample surface a water spray was also included. The laser produced grooves and cuts were analyzed by light microscopy and laser scanning microscopy regarding to the ablation quality, geometry, ablation efficacy, and thermal effects.The grooves in dentin and enamel show a rough surface, typical for Er:YAG laser ablation. The craters are slightly cone shaped with sharp edges on the surface. Water cooling is essential to prevent thermal injury. The ablation efficacy in dentin is comparable to literature values of the flash lamp pumped Er:YAG laser. The cutting of bone and soft tissue is excellent and appears superior to earlier results obtained with flash lamp pumped system. As a further advantage, the broad range of repetition rates allows to widely vary the thermal side effects.In conclusion, these first experiments with a diode pumped Er:YAG laser system on dental hard and soft tissue demonstrate its ability for use in dentistry.