High-throughput and label-free parasitemia quantification and stage differentiation for malaria-infected red blood cells

Abstract

This work reports a high throughput and label-free microfluidic cell deformability sensor for quantitative parasitemia measurement and stage determination for Plasmodium falciparum-infected red blood cells (Pf-iRBCs). The sensor relies on differentiating the RBC deformability (a mechanical biomarker) that is highly correlated with the infection status. The cell deformability is measured by evaluating the transit time when each individual RBC squeezes through a microscale constriction (cross-section ~5μm×5μm). More than 30,000 RBCs can be analyzed for parasitemia quantification in under 1 min with a throughput ~500 cells/s. Moreover, the device can also differentiate various malaria stages (ring, trophozoite, and schizont stage) due to their varied deformability. Using Pf-iRBCs at 0.1% parasitemia as a testing sample, the microfluidic deformability sensor achieved an excellent sensitivity (94.29%), specificity (86.67%) and accuracy (92.00%) in a blind test, comparable to the gold standard of the blood smear microscopy. As a supplement technology to the microscopy and flow cytometry, the microfluidic deformability sensor would possibly allow for label-free, rapid and cost-effective parasitemia quantification and stage determination for malaria in remote regions.

Microfluidic deformability-activated sorting of single particles

“…A buffering region was included to reduce the crosstalk between the consecutive sensing and sorting. The deformability sensing was indirectly performed by the previously validated constriction-based transit time measurement 10,[25][26][27][28] . Briefly, soft particles take less time to squeeze through the sensing pore, while rigid particles take more time (Fig.…”

“…Abnormalities in cell deformability are associated with disease pathogenesis and progression. For instance, metastatic cancer cells are 70% more deformable than benign cells, promoting metastasis [1][2][3] ; the erythrocyte stiffness changes in cytoskeletal disorders such as spherocytosis 4,5 and sickle cell anemia 6,7 as well as in infectious diseases such as malaria [8][9][10] . As a result, deformability has emerged as an intriguing label-free biomarker [11][12][13][14][15][16] .…”

“…Traditional single-particle deformability measurements include micropipette aspiration 23 , optical stretching 24 , atomic force microscopy (AFM) 1 , and magnetic bead-based rheology 2 . To increase the throughput, microfluidic approaches have been developed that rely on either the physical constriction 10,[25][26][27][28] or the hydrodynamic shear stress from a channel [29][30][31] , a cross-section 32 , or a Tjunction 33 .…”

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“…[11] The diagnosis using the microscopy [9] method still has the edge over other techniques for the quantification of the malaria parasites. [11,12] There are several advantages of the microscopic techniques which include the ability to distinguish between the different species of the malaria parasite, quantification of the parasite with sensitivity up to 0.001% and also observation of the asexual stages of the malaria parasites. [13,14] The limitations of the microscopic technique include lack of expertise in analysis of Giemsa-stained blood smears, individualism, chances of non-reproducibility, and also it is time-consuming (it may take >5 min to analyze one slide).…”

“…The principle of this technique is the detection of the various spin-spin relaxation times of the signals observed in MRR forms the hemozoin particles which were paramagnetic. [12] Raman Spectroscopy is a labelfree imaging technique which gives unique and particular Raman fingerprint spectrum of various biological samples. This technique was used to observe what changes took place in the molecular composition of a mice spleen tissue when malaria-infected it as compared to standard or non-infected spleen tissues.…”