Cell refractive index is a key biophysical parameter, which has been extensively studied. It is correlated with other cell biophysical properties including mechanical, electrical and optical properties, and not only represents the intracellular mass and concentration of a cell, but also provides important insight for various biological models. Measurement techniques developed earlier only measure the effective refractive index of a cell or a cell suspension, providing only limited information on cell refractive index and hence hindering its in-depth analysis and correlation. Recently, the emergence of microfluidic, photonic and imaging technologies has enabled the manipulation of a single cell and the 3D refractive index of a single cell down to sub-micron resolution, providing powerful tools to study cells based on refractive index. In this review, we provide an overview of cell refractive index models and measurement techniques including microfluidic chip-based techniques for the last 50 years, present the applications and significance of cell refractive index in cell biology, hematology, and pathology, and discuss future research trends in the field, including 3D imaging methods, integration with microfluidics and potential applications in new and breakthrough research areas.

Cell refractive index for cell biology and disease diagnosis: past, present and future

The past two decades have witnessed the revolutionary development of optical trapping of nanoparticles, most of which deal with trapping stiffness larger than 10-8 N/m. In this conventional regime, however, it remains a formidable challenge to sort out sub-50-nm nanoparticles with single-nanometer precision, isolating us from a rich flatland with advanced applications of micromanipulation. With an insightfully established roadmap of damping, the synchronization between optical force and flow drag force can be coordinated to attempt the loosely overdamped realm (stiffness, 10-10 to 10-8 N/m), which has been challenging. This paper intuitively demonstrates the remarkable functionality to sort out single gold nanoparticles with radii ranging from 30 to 50 nm, as well as 100- and 150-nm polystyrene nanoparticles, with single nanometer precision. The quasi-Bessel optical profile and the loosely overdamped potential wells in the microchannel enable those aforementioned nanoparticles to be separated, positioned, and microscopically oscillated. This work reveals an unprecedentedly meaningful damping scenario that enriches our fundamental understanding of particle kinetics in intriguing optical systems, and offers new opportunities for tumor targeting, intracellular imaging, and sorting small particles such as viruses and DNA.

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Nanometer-precision linear sorting with synchronized optofluidic dual barriers

The feasibility of identifying cancer cells by measuring the refractive index (RI) distribution across a single live cell with ultrahigh resolution full-field optical coherence microscopy (FF-OCM) is presented. The FF-OCM is utilized to quantify integral RI distributions of unmodified cells without any cell treatments and used as a biophysical indicator for diagnosing cell malignancy. Firstly, the physical thickness distribution of the cell adherent to a culture dish is measured by taking a series of 0.6 µm resolved en-face tomograms. Subsequently, from the en-face image of the bottom surface of the cell or the top surface of the dish, the phase gain image of the cell is extracted. Then, from these two measurements the axially averaged RI map of the cell is extracted. The implemented FF-OCM system had a 0.8 µm axial resolution and the phase measurement sensitivity of the system was around 124 mrad. With the system, RI maps of several living cell lines of normal and cancer cells were constructed and quantitatively analyzed. The experiments showed that cancer cells had higher RI than normal ones. This approach using the FF-OCM has significant potential for cancer diagnosis and dynamic cell analysis as in situ label-free biophysical assay.

Full-field optical coherence microscopy for identifying live cancer cells by quantitative measurement of refractive index distribution

This critical review summarizes the developments in the integration of micro-optical elements with microfluidic platforms for facilitating detection and automation of bio-analytical applications. Micro-optical elements, made by a variety of microfabrication techniques, advantageously contribute to the performance of an analytical system, especially when the latter has microfluidic features. Indeed the easy integration of optical control and detection modules with microfluidic technology helps to bridge the gap between the macroscopic world and chip-based analysis, paving the way for automated and high-throughput applications. In our review, we start the discussion with an introduction of microfluidic systems and micro-optical components, as well as aspects of their integration. We continue with a detailed description of different microfluidic and micro-optics technologies and their applications, with an emphasis on the realization of optical waveguides and microlenses. The review continues with specific sections highlighting the advantages of integrated micro-optical components in microfluidic systems for tackling a variety of analytical problems, like cytometry, nucleic acid and protein detection, cell biology, and chemical analysis applications.

Micro-optics for microfluidic analytical applications

The integration of microactuators within a silicon photonic chip gave rise to the field of optical micro-electro-mechanical systems (MEMS) that was originally driven by the telecommunication market. Following the latter's bubble collapse in the beginning of the third millennium, new directions of research with considerable momentum appeared focusing on the realization and applications of miniaturized instrumentation in biology, chemistry, physics and materials science. At the heart of these applications light interferometry is a key optical phenomenon, in which miniaturized scanning interferometers are the manipulating optical devices. Monolithic free-space optical interferometers realized on a silicon chip take advantage of the recent progress in the microfabrication technology that is enabling accurate control of the etching depth, the aspect ratio, the verticality and the curvature of the etched surfaces. The fabrication technology, the library of micro-optical and mechanical components, the realized architectures and their characterization are described in detail in this review, followed by a discussion of the foreseen challenges.

Monolithic silicon‐micromachined free‐space optical interferometers onchip

This letter reports the measurement of single living cell’s refractive index (RI) using the optical grating resonant cavity with buffer modulation on a biochip. The cell’s RI is a significant cell biophysical property which has potential in cancer and disease diagnoses. The single cell is trapped optically within the cavity, and the transmission spectra shift due to the change of surrounding buffer, without/with cell, are used to determine the cell’s effective RI and effective thickness. The measurement shows that Madin-Darby canine kidney cell has an effective RI of 1.383±0.001. Moreover, the optical trapping technique eliminates uncertainty and avoids cell damage.

https://nocweba.ntu.edu.sg/laq_mems/journal%20papers/2007/Differential%20single%20living%20cell%20refractometry%20using%20grating%20resonant%20cavity%20with%20optical%20trap.pdf

S. Takahashi

Papers

Differential single living cell refractometry using grating resonant cavity with optical trap