Breast cancer is most common cancer among women world-wide and can be treated if diagnosed at early-stage. Fluorescence (FL) techniques have a considerable impact in tumor detection as FL is highly sensitive to biochemical and biophysical structure of the tissues, providing novel techniques for early and noninvasive diagnosis of cancer. Due to high sensitivity of FL, it can be used in early-stage breast cancer detection. Fluorescence-guided-surgery (FGS) using exogenous agent in breast cancer is a well-known method, which locates tumor and margins during intra-operative procedure. FL-spectroscopy and FL-imaging has shown the potentiality in FGS of breast cancer independently. Combination of these two modalities in a single system improves the success rate of FGS which can give tumor free tissues. The goal of the current study is to analyze and classify the early-stage breast cancer, locally advanced breast cancer (invasive ductal carcinoma) and normal tissue. For that, we combine FL-imaging and FL-spectroscopy in a single smartphone-based point-of-care devices and recorded data during intra-operative procedure. A total 21 patients of invasive ductal carcinoma and fibroadenoma are included in this study. Total 65 FL-spectra are recorded during intra-operative procedure which are further used in support vector machine (SVM) based classification of fibroadenoma, invasive ductal carcinoma (IDC) and normal tissue. The sensitivity, specificity, positive predictive value, negative predictive value, and overall diagnostic accuracy (total efficiency) is 78.6 %, 90%, 91.6%, 75%, 95%, respectively. Additionally, we observed a red shift in case of IDC and fibroadenoma from normal tissue, which is 5.22±1.77 nm, and 4.96±2.61 nm, respectively.
We describe field-portable GRIN lens based micro-endoscope with oblique-illumination for cancer screening. Fluorescence microscopic images of different samples were recorded with micro-endoscope which provides molecular information about the sample.
We report classification between normal and anemic erythrocytes by determining cell counts computationally using Circular Hough transform algorithm in matlab and quantifying phase map which are important for early diagnosis of diseases.
We demonstrate a multi-modal system to obtain diverse information about biological specimen in single-shot. The quantitative analysis of MG63 osteosarcoma cells is presented which are cultured on Si substrate and stained using sodium fluorescein dye.
We report the development of field-portable multi-modal chip-based fluorescence, bright field and quantitative phase microscopy using smartphone detecting system. Fluorescence microscopy provide molecular information of the specimen with excellent specificity, while phase microscopy provides quantitative information of the specimen. Quantifying the optical phase shifts associated with biological structures gives access to information about morphology and dynamics at the nanometer scale. Here, we propose an integrated waveguide chip-based total internal reflection fluorescence (TIRF) microscopy and quantitative phase microscopy (QPM). We have developed microLED with cylindrical beam profile to couple excitation light into the edges of glass slide easily and efficiently. The evanescent field present on top of a waveguide surface is used to excite the fluorescence and a mobile phone microscope is used to collect the signal. Waveguide chip-based TIRF microscopy benefits from decoupling of illumination and collection light path, large field of view imaging and pre-aligned configuration for multi-color TIRF imaging. Light for bright field imaging and QPM integrated in the transmission mode. A microscope objective is used for collecting the fluorescence excited by evanescent field and transmitted light for bright field and quantitative phase microscopy (QPM). A compact and common path interferometer is used for QPM. The entire device is fabricated using three-D printer and integrated into one, which is compact and field portable. Images are recorded using a smart phone. Experimental results of onion epithelial cells, polystyrene microspheres and normal breast tissue are presented. The cost of entire system is very less.
Quantitative phase microscopy (QPM) is a label-free imaging technique to quantify various biophysical parameters, such as refractive index, optical thickness, cell dry mass, and dynamic membrane fluctuations. Accurate determination of these parameters requires the use of a QPM system with high temporal phase stability and high spatial phase sensitivity. We report a QPM system based on a common-path interferometer with high temporal phase stability and high spatial phase sensitivity. The proposed QPM system is highly temporally stable, compact and easy to align and implement. The interference pattern can be obtained quickly even with a low coherent light source. In order to realize high spatial phase sensitivity, we used partially spatially coherent (pseudo-thermal) light source for illumination. Due to the partial spatial coherent nature of the light source, a speckle-free interferogram/hologram is recorded over the entire field-of-view. Two types of speckle free QPM systems are implemented using common path Fresnel biprism as well as lateral shearing interferometers. A Fresnel biprism is used in the self-referencing mode, thus offering the advantage of no optical power loss in addition to high temporal stability and the least speckle artifacts. Furthermore, it is very easy to implement, as the system completely replaces the need for spatial filtering at the source end as well as for the reference beam generation. In another configuration, we used a lateral shearing interferometer. The scattered light from the object is collected by the microscope objective lens and passes through a 4mm thick optically flat parallel plate to generate the interference pattern. Phase maps of human RBCs are reconstructed and the results are compared for fully and partially coherent light illumination.
We report longitudinal spatial coherence (LSC) gated high-resolution tomography and quantitative phase microscopy of biological cells and tissues with uniform illumination using laser as a light source. To accomplish this a pseudo thermal light source was synthesized by passing laser beams through an optical system, which is basically a speckle reduction system with combined effect of spatial, temporal, angular and polarisation diversity. The longitudinal spatial coherence length of such light was significantly reduced by synthesizing a pseudo thermal source with the combined effect of spatial, angular and temporal diversity. This results in a low spatially coherent (i.e., broad angular frequency spectrum) light source with narrow temporal frequency spectrum. Light from such a pseudo thermal light source was passed through an interference microscope with varying magnification, such as, 10X and 50X. The interference microscope was used for full-field OCT imaging of multilayer objects and topography of industrial objects. Experimental results of optical sectioning of multilayer biological objects with high axial-resolution less than 10μm was achieved which is comparable to broadband white light source. The synthesized light source with reduced speckles having uniform illumination on the sample, which can be very useful for fluorescence microscopy as well as quantitative phase microscopy with less phase noise. The present system does not require any dispersion compensation optical system for biological samples as a highly monochromatic light source is used.
In digital holographic interferometry (DHI), coherent noise degrades accuracy of phase
information. We present multi-beam polarization DHI in which two cross polarized interferograms are
recorded. Fourier analysis of interferograms reduces coherent noise and increases accuracy.
A spectrally resolved white light interferometry is demonstrated using a discrete
spectrum light sources (i.e., RGB LEDs) and monochrome CCD camera for the multi-color
quantitative phase imaging of biological cells without color cross talk.
We demonstrate results for phase maps of biological cells using white-light and multi-spectral
interference microscopy. Study on comparison of phase maps reconstructed using 1-CCD and 3-
CCD is presented to reduce color cross-talk and improved resolution.
A spatially low coherent light source is synthesized to reduce the spatial phase noise in
the laser based digital holographic microscope, which otherwise introduces unwanted spatial
phase, subsequently, the height measurement error of the biological objects.
We present a single-shot white light interference microscopy for the quantitative phase imaging (QPI) of biological cells and tissues. A common path white light interference microscope is developed and colorful white light interferogram is recorded by three-chip color CCD camera. The recorded white light interferogram is decomposed into the red, green and blue color wavelength component interferograms and processed it to find out the RI for different color wavelengths. The decomposed interferograms are analyzed using local model fitting (LMF)" algorithm developed for reconstructing the phase map from single interferogram. LMF is slightly off-axis interferometric QPI method which is a single-shot method that employs only a single image, so it is fast and accurate. The present method is very useful for dynamic process where path-length changes at millisecond level. From the single interferogram a wavelength-dependent quantitative phase imaging of human red blood cells (RBCs) are reconstructed and refractive index is determined. The LMF algorithm is simple to implement and is efficient in computation. The results are compared with the conventional phase shifting interferometry and Hilbert transform techniques.
This paper describes a new and novel phase shifting technique for qualitative as well as quantitative measurement in microscopy. We have developed a phase shifting device which is robust, inexpensive and involves no mechanical movement. In this method, phase shifting is implemented using LED array, beam splitters and defocused projection of Ronchi grating. The light from the LEDs are made incident on the beam splitters at spatially different locations. Due to variation in the geometrical distances of LEDs from the Ronchi grating and by sequentially illuminating the grating by switching on one LED at a time the phase shifted grating patterns are generated. The phase shifted structured patterns are projected onto the sample using microscopic objective lens. The phase shifted deformed patterns are recorded by a CCD camera. The initial alignment of the setup involves a simple procedure for the calibration for equal fringe width and intensity such that the phase shifted fringes are at equal phase difference. Three frame phase shifting algorithm is employed for the reconstruction of the phase map. The method described here is fully automated so that the phase shifted images are recorded just by switching of LEDs and has been used for the shape measurement of microscopic industrial objects. The analysis of the phase shifted images provides qualitative as well as quantitative information about the sample. Thus, the method is simple, robust and low cost compared to PZT devices commonly employed for phase shifting.
We report white light phase shifting interferometry in conjunction with color fringe analysis for the detection of contaminants in water such as Escherichia coli (E.coli), Campylobacter coli and Bacillus cereus. The experimental setup is based on a common path interferometer using Mirau interferometric objective lens. White light interferograms are recorded using a 3-chip color CCD camera based on prism technology. The 3-chip color camera have lesser color cross talk and better spatial resolution in comparison to single chip CCD camera. A piezo-electric transducer (PZT) phase shifter is fixed with the Mirau objective and they are attached with a conventional microscope. Five phase shifted white light interferograms are recorded by the 3-chip color CCD camera and each phase shifted interferogram is decomposed into the red, green and blue constituent colors, thus making three sets of five phase shifted intererograms for three different colors from a single set of white light interferogram. This makes the system less time consuming and have lesser effect due to surrounding environment. Initially 3D phase maps of the bacteria are reconstructed for red, green and blue wavelengths from these interferograms using MATLAB, from these phase maps we determines the refractive index (RI) of the bacteria. Experimental results of 3D shape measurement and RI at multiple wavelengths will be presented. These results might find applications for detection of contaminants in water without using any chemical processing and fluorescent dyes.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.