On-chip flow cytometer using integrated photonics for the detection of human leukocytes.

Differentiation between leukocyte subtypes like monocytes and lymphocytes is essential for cell therapy and research applications. To guarantee the cost-effective delivery of functional cells in cell therapies, billions of cells must be processed in a limited time. Yet, the sorting rates of commercial cell sorters are not high enough to reach the required yield. Process parallelization by using multiple instruments increases variability and production cost. A compact solution with higher throughput can be provided by multichannel flow cytometers combining fluidics and optics on-chip. In this work, we present a micro-flow cytometer with monolithically integrated photonics and fluidics and demonstrate that both the illumination of cells, as well as the collection of scattered light, can be realized using photonic integrated circuits. Our device is the first with sufficient resolution for the discrimination of lymphocytes and monocytes. Innovations in microfabrication have enabled complete integration of miniaturized photonic components and fluidics in a CMOS-compatible wafer stack. In combination with external optics, the device is ready for the collection of fluorescence using the on-chip excitation.

Ultra-compact silicon photonic integrated interferometer for swept-source optical coherence tomography.

We demonstrate an ultra-compact silicon integrated photonic interferometer for swept-source optical coherence tomography (SS-OCT). The footprint of the integrated interferometer is only 0.75×5 mm2. The design consists of three 2×2 splitters, a 13 cm physical length (50.4 cm optical length) reference arm, and grating couplers. The photonic integrated circuit was used as the interferometer of an SS-OCT system. The sensitivity of the system was measured to be -62 dB with 115 µW power delivered to the sample. Using the system, we demonstrate cross-sectional OCT imaging of a layered tissue phantom. We also discuss potential improvements in passive silicon photonic integrated circuit design and integration with active components.

Photonic integrated Mach-Zehnder interferometer with an on-chip reference arm for optical coherence tomography.

Optical coherence tomography (OCT) is a noninvasive, three-dimensional imaging modality with several medical and industrial applications. Integrated photonics has the potential to enable mass production of OCT devices to significantly reduce size and cost, which can increase its use in established fields as well as enable new applications. Using silicon nitride (Si3N4) and silicon dioxide (SiO2) waveguides, we fabricated an integrated interferometer for spectrometer-based OCT. The integrated photonic circuit consists of four splitters and a 190 mm long reference arm with a foot-print of only 10 × 33 mm(2). It is used as the core of a spectral domain OCT system consisting of a superluminescent diode centered at 1320 nm with 100 nm bandwidth, a spectrometer with 1024 channels, and an x-y scanner. The sensitivity of the system was measured at 0.25 mm depth to be 65 dB with 0.1 mW on the sample. Using the system, we imaged human skin in vivo. With further optimization in design and fabrication technology, Si3N4/SiO2 waveguides have a potential to serve as a platform for passive photonic integrated circuits for OCT.

Heterodyne laser Doppler vibrometers integrated on silicon-on-insulator based on serrodyne thermo-optic frequency shifters.

Miniaturized laser Doppler vibrometers (LDVs) have many advantages over conventional bulk LDVs. In this paper, the realization of a miniaturized heterodyne LDV integrated on silicon-on-insulator substrate is reported. The optical frequency shifters in these on-chip LDVs employ a serrodyne technique, and they generate a frequency shift at 2 kHz. Vibrations of a mirror for the frequency range between 1.1 and 123 Hz and the velocity range between 0.8 and 400 µm/s are measured by both an on-chip LDV and a commercial LDV. The measurement results agree well. A compensation method for the influence of on-chip spurious reflections is also demonstrated.

Compact grating couplers on silicon-on-insulator with reduced backreflection.

The backreflection in commonly used grating couplers on silicon-on-insulator (SOI) is not negligible for many applications. This reflection is dramatically reduced in our improved compact grating coupler design, which directs the reflection away from the input waveguide. Realized devices on SOI show that the reflection can be reduced down to -50 dB without an apparent transmission penalty.

Rapid pulse shaping with homodyne detection for measuring nonlinear optical signals.

We have designed a common-mode interferometric acousto-optic pulse shaper that is capable of shaping individual pulses differently from a mode-locked laser. The design enables the measurement of weak nonlinear optical signals such as two-photon absorption and self-phase modulation at megahertz rates. The experimental apparatus incorporates homodyne detection as a means of resolving the phase of the detected signals. The fast data acquisition rate and the ability to perform measurements in scattering media make this experimental apparatus amenable to imaging applications analogous to measurements of two-photon fluorescence using a mode-locked laser.

Two-color, two-photon, and excited-state absorption microscopy.

We develop a new approach in imaging nonfluorescent species with two-color two-photon and excited state absorption microscopy. If one of two synchronized mode-locked pulse trains at different colors is intensity modulated, the modulation transfers to the other pulse train when nonlinear absorption takes places in the medium. We can easily measure 10(-6) absorption changes caused by either two-photon absorption or excited-state absorption with a RF lock-in amplifier. Sepia melanin is studied in detail as a model system. Spectroscopy studies on the instantaneous two-photon absorption (TPA) and the relatively long-lived excited-state absorption (ESA) of melanin are carried out in solution, and imaging capability is demonstrated in B16 cells. It is found that sepia melanin exhibits two distinct excited states with different lifetimes (one at 3 ps, one lasting hundreds of nanoseconds) when pumped at 775 nm. Its characteristic TPA/ESA enables us to image its distribution in cell samples with high resolution comparable to two-photon fluorescence microscopy (TPFM). This new technique could potentially provide valuable information in diagnosing melanoma.

Two-photon absorption and self-phase modulation measurements with shaped femtosecond laser pulses.

We show that phase-sensitive detection of spectral hole refilling can yield information about self-phase modulation and two-photon absorption coefficients. We expect that, when applied to tissue microscopy, this technique will allow the study of endogenous molecular markers beneath the surface, even if those markers are nonfluorescent.

Mitochondrial NADH as the bellwether of tissue O2 delivery.

It is proposed that the redox state of mitochondrial NADH will complement blood gas analysis for measuring the health and welfare of human tissues. Use of arterial oxygen saturation levels (SaO2), especially as assayed by the Nellcor instrument, has spread almost everywhere in medicine despite the fact that hypoxia of internal organs, liver, kidney, brain, pancreas, etc. is not well indicated by peripheral digital oxygenation. Indeed, there is an implied liability in the failure to infer central oxygenation from peripheral values. Near infrared (NIR) sensing of deep tissue saturation of hemoglobin (StO2) requires multi-wavelength, multi-site measurement of both absorption and scattering properties by time or frequency domain NIR methods. Corrections for underlying water and lipid absorptions can be made so that the correct value for, and saturation oh hemoglobin are obtained. Nevertheless, the significance of blood oxygen saturation, even localized to particular organs, can be questioned from the standpoint of what is the critical value of the desaturation from which the tissue can recover; for example, in the case of cortical neurons where stroke, compression ischemia, etc. cause O2 lack, this value becomes of significant clinical importance in both the brain and the spinal chord. These approaches are actively pursued and the possibility of subsurface redox state measurement in human tissues may eventually emerge as the quantitative metric of tissue metabolic state and of hypoxic stress. The great flexibility and versatility of the fast, economical and "tetherless" nature of opto-electronic technology is appropriate to the manifold challenges of neuronal function as currently measured by intrinsic signals and soon to be studiable by extrinsic signals of metabolism and electrophysiological functions.