C. Dongre

Three-dimensional Mach-Zehnder interferometer in a microfluidic chip for spatially-resolved label-free detection

Ultrafast laser writing of waveguides in glasses is a very flexible and simple method for direct on-chip integration of photonic devices. In this work we present a monolithic optofluidic device in fused silica providing label-free and spatially-resolved sensing in a microfluidic channel. A Mach-Zehnder interferometer is inscribed with the sensing arm orthogonally crossing the microfluidic channel and the reference arm passing over it. The interferometer is integrated either with a microchannel fabricated by femtosecond laser technology or into a commercial lab-on-chip for capillary electrophoresis. The device layout, made possible by the unique three-dimensional capabilities of the technique, enables label-free sensing of samples flowing in the microchannel with spatial resolution of about 10 microm and limit of detection down to 10(-4) RIU.

Fluorescence monitoring of microchip capillary electrophoresis separation with monolithically integrated waveguides

Using femtosecond laser writing, optical waveguides were monolithically integrated into a commercial microfluidic lab-on-a-chip device, with the waveguides intersecting a microfluidic channel. Continuous-wave laser excitation through these optical waveguides confines the excitation window to a width of 12 microm, enabling high-resolution monitoring of the passage of different types of fluorescent analytes when migrating and being separated in the microfluidic channel by microchip capillary electrophoresis. Furthermore, we demonstrate on-chip-integrated waveguide excitation and detection of a biologically relevant species, fluorescently labeled DNA molecules, during microchip capillary electrophoresis. Well-controlled plug formation as required for on-chip integrated capillary electrophoresis separation of DNA molecules, and the combination of waveguide excitation and a low limit of detection, will enable monitoring of extremely small quantities with high spatial resolution.

Optical sensing in microfluidic lab-on-a-chip by femtosecond-laser-written waveguides

AbstractWe use direct femtosecond laser writing to integrate optical waveguides into a commercial fused silica capillary electrophoresis chip. High-quality waveguides crossing the microfluidic channels are fabricated and used to optically address, with high spatial selectivity, their content. Fluorescence from the optically excited volume is efficiently collected at a 90° angle by a high numerical aperture fiber, resulting in a highly compact and portable device. To test the platform we performed electrophoresis and detection of a 23-mer oligonucleotide plug. Our approach is quite powerful because it allows the integration of photonic functionalities, by simple post-processing, into commercial LOCs fabricated with standard techniques. FigureFemtosecond laser written waveguides can selectively excite fluorescence in a microfluidic channel of a commercial lab-on-a-chip. A compact scheme for on-chip detection by laser induced fluorescence is applied to capillary electrophoresis of a 23-mer Cy3-labeled oligonucleotide

High-resolution electrophoretic separation and integrated-waveguide excitation of fluorescent DNA molecules in a lab on a chip

By applying integrated‐waveguide laser excitation to an optofluidic chip, fluorescently labeled DNA molecules of 12 or 17 different sizes are separated by CE with high operating speed and low sample consumption of ∼600 pL. When detecting the fluorescence signals of migrating DNA molecules with a PMT, the LOD is as low as 2.1 pM. In the diagnostically relevant size range (∼150–1000 base‐pairs) the molecules are separated with reproducibly high sizing accuracy (>99%) and the plug broadening follows Poissonian statistics. Variation of the power dependence of migration time on base‐pair size – probably with temperature and condition of the sieving gel matrix – indicates that the capillary migration cannot be described by a simple physical law. Integrated‐waveguide excitation of a 12‐μm narrow microfluidic segment provides a spatio‐temporal resolution that would, in principle, allow for a 20‐fold better accuracy than the currently supported by state‐of‐the‐art electrophoretic separation in microchips, thereby demonstrating the potential of this integrated optical approach to fulfill the resolution demands of future electrophoretic microchips.

Dual-point dual-wavelength fluorescence monitoring of DNA separation in a lab on a chip.

We present a simple approach in electrophoretic DNA separation and fluorescent monitoring that allows to identify the insertion or deletion of base-pairs in DNA probe molecules from genetic samples, and to perform intrinsic calibration/referencing for highly accurate DNA analysis. The principle is based on dual-point, dual-wavelength laser-induced fluorescence excitation using one or two excitation windows at the intersection of integrated waveguides and microfluidic channels in an optofluidic chip and a single, color-blind photodetector, resulting in a limit of detection of ~200 pM for single-end-labeled DNA molecules. The approach using a single excitation window is demonstrated experimentally, while the option exploiting two excitation windows is proposed theoretically.

All-numerical noise filtering of fluorescence signals for achieving ultra-low limit of detection in biomedical applications

We present an all-numerical method for post-processing of the fluorescent signal as obtained from labeled molecules by capillary electrophoresis (CE) in an optofluidic chip, on the basis of data filtering in the Fourier domain. It is shown that the method outperforms the well-known lock-in amplification during experiments in the reduction of noise by a factor of (square root)2. The method is illustrated using experimental data obtained during CE separation of molecules from a commercial DNA ladder with 17 fluorescently labeled molecules having different base-pair sizes. An improvement in signal-to-noise ratio by a factor of ∼10 is achieved, resulting in a record-low limit of detection of 210 fM.

Femtosecond laser fabrication for the integration of optical sensors in microfluidic lab-on-chip devices

Femtosecond lasers enable the fabrication of both optical waveguides and buried microfluidic channels on a glass substrate. The waveguides are used to integrate optical detection in a commercial microfluidic lab-on-chip for capillary electrophoresis.

Electrophoretic Separation and Detection of a Few DNA Molecules in An Optofluidic Chip

After electrophoretic separation of dye-labeled DNA molecules of 17 different sizes, integrated-waveguide laser excitation and physical or numerical lock-in amplification enables a limit of detection down to 8-9 DNA molecules in an optofluidic chip.

Fluorescence monitoring of capilarry electrophoresis separation in a lab-on-a-chip with monolithically integrated waveguides

The operation dynamics of end-pumped solid-state lasers are investigated by means of a spatially resolved numerical rate-equation model and a time-dependent analytical thermal model. The rate-equation model allows the optimization of parameters such as the output coupler transmission and gain medium length, with the aim of improving the laser output performance. The time-dependent analytical thermal model is able to predict the temperature and the corresponding induced thermal stresses on the pump face of quasi-continuous wave (qcw) end-pumped laser rods. Both models are found to be in very good agreement with experimental results.

Combined microfluidic-optical DNA analysis with single-base-pair sizing capability

DNA sequencing by microchip capillary electrophoresis (CE) enables cheap, high-speed analysis of low reagent volumes. One of its potential applications is the identification of genomic deletions or insertions associated with genetic illnesses. Detecting single base-pair insertions or deletions from DNA fragments in the diagnostically relevant size range of 150-1000 base-pairs requires a variance of σ2 < 10-3. In a microfluidic chip post-processed by femtosecond-laser writing of an optical waveguide we CE-separated 12 blue-labeled and 23 red-labeled DNA fragments in size. Each set was excited by either of two lasers power-modulated at different frequencies, their fluorescence detected by a photomultiplier, and blue and red signals distinguished by Fourier analysis. We tested different calibration strategies. Choice of the fluorescent label as well as the applied fit function strongly influence the obtained variance, whereas fluctuations between two consecutive experiments are less detrimental in a laboratory environment. We demonstrate a variance of σ2 ≈4 × 10-4, lower than required for the detection of single base-pair insertion or deletion in an optofluidic chip.