Development of Quantitative Phase Imaging as a Diagnostic Modality with High-Throughput Implementation

Without exogenous contrast agents, imaging semitransparent samples such as biological cells can be difficult with light microscopy that measures modulation in the intensity of transmitted light. Quantitative phase microscopy (QPM) that measures the phase delays imparted by the objects is often used to examine the spatial and temporal dynamics of individual cells without chemical modification.

In order to demonstrate its effectiveness as a diagnostic tool, quantitative phase microscope is used to image red blood cells (RBCs) specifically those infected with malaria parasites, Plasmodium falciparum. RBCs are a favorite target for QPM studies, in particular because they have little internal structure and thus can be represented by a homogenous refractive index. The ability to profile RBCs is an important capability of QPM since these cells are affected by different disease stages, and thus they are essential in human health diagnostics. Classifiers built using QPM are able to distinguish and classify uninfected red blood cells from those infected with P. falciparum using morphological features of the cells with high accuracy. The study shows that QPM can be a very useful diagnostic modality and that it can be more clinically relevant by developing into a high throughput holographic imaging system.

In addition to the morphological measurement of erythrocytes, biophysical properties of RBCs under mechanical stress are measured by incorporating microfluidic chips into the QPM platform. Highly precise microfluidic chips are manufactured with specific dimensions that will stress the RBCs at designated positions as they flow through them. Using a high refractive index medium, QPM is used to measure the optical volume changes associated with efflux of water through the membrane of the cells under mechanical stress. The OV changes in response to mechanical force are compared for different storage periods to evaluate the changes in response to deformation throughout the ageing cells.

Finally, a novel approach for high throughput screening is developed based on holographic imaging of the cells flowing through microfluidic chips. To enable high throughput imaging, we have implemented a new quantitative phase imaging modality, holographic cytometry. Holographic cytometry maintains the high sensitivity of QPM while imaging a large number of cells flowing through the microfluidic devices. As demonstrated in previous studies, morphological parameters are extracted from these images to assess their changes over the storage time and classify them according to the time in the blood units.