a-rapid-chemical-free-hemoglobin-assay-with-photothermal-angular

A Rapid and Chemical-free Hemoglobin Assay with Photothermal Angular Light Scattering

Photo-thermal angular light scattering (PT-AS) is a novel optical method for measuring the hemoglobin concentration ([Hb]) of blood samples. On the basis ...

A novel optical detection method for hemoglobin concentration is described. The hemoglobin molecules consisting mainly of iron generate heat upon their absorption of light energy at 532 nm, which subsequently changes the refractive index of the blood. We exploit this photothermal effect to determine the hemoglobin concentration of erythrocytes without any preprocessing of blood. Highly sensitive measurement of refractive index alteration of blood samples is enabled by a spectral-domain low coherence reflectometric sensor with subnanometer-level optical path-length sensitivity. The performance and validity of the sensor are presented by comparing the measured results against the reference data acquired from an automatic hematology analyzer.

Photothermal spectral-domain optical coherence reflectometry for direct measurement of hemoglobin concentration of erythrocytes.

Quantitative measurement of dynamic responses of unstained living cells is of great importance in many applications ranging from investigation of fundamental cellular functions to drug discoveries. Conventional optical methods for label-free cell-based assay examine cellular structural changes proximal to sensor surfaces under external stimuli, but require dedicated nanostructure-patterned substrates for operation. Here, we present a quantitative imaging method, spectral-domain optical coherence phase microscopy (SD-OCPM), as a viable optical platform for label-free cell-based assay. The instrument is based on a low-coherence interferometric microscope that enables quantitative depth-resolved phase measurement of a transparent specimen with high phase stability. We demonstrate SD-OCPM measurement of dynamic responses of human breast cancer cells (MCF-7) to 2-picolinic acid (PA) and histamine.

Label-free cell-based assay with spectral-domain optical coherence phase microscopy.

We present an implementation of spectrally encoded slit confocal microscopy. The method employs a rapid wavelength-swept laser as the light source and illuminates a specimen with a line focus that scans through the specimen as the wavelength sweeps. The reflected light from the specimen is imaged with a stationary line scan camera, in which the finite pixel height serves as a slit aperture. This scanner-free operation enables a simple and cost-effective implementation in a small form factor, while allowing for the three-dimensional imaging of biological samples.

Spectrally encoded slit confocal microscopy using a wavelength-swept laser.

We describe a three-dimensional microscopy technique based on spectral and frequency encoding. The method employs a wavelength-swept laser to illuminate a specimen with a spectrally-dispersed line focus that sweeps over the specimen in time. The spatial information along each spectral line is further mapped into different modulation frequencies. Spectrally-resolved detection and subsequent Fourier analysis of the back-scattered light from the specimen therefore enable high-speed, scanner-free imaging of the specimen with a single-element photodetector. High-contrast, three-dimensional imaging capability of this method is demonstrated by presenting images of various materials and biological specimens.

Frequency- and spectrally-encoded confocal microscopy.

We present a direct, rapid and chemical-free detection method for hemoglobin concentration ([Hb]), based on photothermal angular light scattering. The iron oxides contained in hemoglobin molecules exhibit high absorption of 532-nm light and generate heat under the illumination of 532-nm light, which subsequently alters the refractive index of blood. We measured this photothermal change in refractive index by employing angular light scattering spectroscopy with the goal of quantifying [Hb] in blood samples. Highly sensitive [Hb] measurement of blood samples was performed by monitoring the shifts in angularly dispersed scattering patterns from the blood-loaded microcapillary tubes. Our system measured [Hb] over the range of 0.35-17.9 g/dL with a detection limit of ~0.12 g/dL. Our sensor was characterized by excellent correlation with a reference hematology analyzer (r>0.96), and yielded a precision of 0.63 g/dL for a blood sample of 9.0 g/dL.

Capillary-scale direct measurement of hemoglobin concentration of erythrocytes using photothermal angular light scattering.

We present a multi-contrast microscope based on color-coded illumination and computation. A programmable three-color light-emitting diode (LED) array illuminates a specimen, in which each color corresponds to a different illumination angle. A single color image sensor records light transmitted through the specimen, and images at each color channel are then separated and utilized to obtain bright-field, dark-field, and differential phase contrast (DPC) images simultaneously. Quantitative phase imaging is also achieved based on DPC images acquired with two different LED illumination patterns. The multi-contrast and quantitative phase imaging capabilities of our method are demonstrated by presenting images of various transparent biological samples.

Article	Year
Photothermal spectral-domain optical coherence reflectometry for direct measurement of hemoglobin concentration of erythrocytes. Biosensors & bioelectronics\| PubMed ID: 24549136	2014
Label-free cell-based assay with spectral-domain optical coherence phase microscopy. Journal of biomedical optics\| PubMed ID: 24711152	2014
Spectrally encoded slit confocal microscopy using a wavelength-swept laser. Journal of biomedical optics\| PubMed ID: 25813913	2015
Frequency- and spectrally-encoded confocal microscopy. Optics express\| PubMed ID: 25836810	2015
Capillary-scale direct measurement of hemoglobin concentration of erythrocytes using photothermal angular light scattering. Biosensors & bioelectronics\| PubMed ID: 26176206	2015
Color-coded LED microscopy for multi-contrast and quantitative phase-gradient imaging. Biomedical optics express\| PubMed ID: 26713205	2015

Suho Ryu

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