Label-free, high-throughput detection ofP. falciparuminfection in sphered erythrocytes with digital holographic microscopy

Abstract

Effective malaria treatment requires rapid and accurate diagnosis of infecting species and actual parasitemia. Despite the recent success of rapid tests, the analysis of thick and thin blood smears remains the gold standard for routine malaria diagnosis in endemic areas. For non-endemic regions, sample preparation and analysis of blood smears are an issue due to low microscopy expertise and few cases of imported malaria. Automation of microscopy results could be beneficial to quickly confirm suspected infections in such conditions. Here, we present a label-free, high-throughput method for early malaria detection with the potential to reduce inter-observer variation by reducing sample preparation and analysis effort. We used differential digital holographic microscopy in combination with two-dimensional hydrodynamic focusing for the label-free detection of P. falciparum infection in sphered erythrocytes, with a parasitemia detection limit of 0.01%. Moreover, the achieved differentiation of P. falciparum ring-, trophozoite- and schizont life cycle stages in synchronized cultures demonstrates the potential for future discrimination of even malaria species.

Image-Based Single Cell Sorting Automation in Droplet Microfluidics

“…The presented system will not be able to compete with high-throughput sorting techniques such as fluorescent activated cell sorting as the full image takes longer to acquire and process, however, it can be used as a complementary technique when small cell populations need to be analysed or when labelling is undesired or when the image data presents a unique opportunity for sorting cell populations. The presented platform could be strengthened by additional sensing modules to aid in the decision making such as differential detection photothermal interferometry 83 , digital holographic microscopy 84 , whispering gallery mode resonators 85 , refractive index cytometry 86 or impedance cytometry 87 .…”

Real-time cholesterol sorting in Plasmodium falciparum-erythrocytes as revealed by 3D label-free imaging

“…A variety of microscopic observations have yielded important information on the biological processes that involve structural modifications inside cells, and recently, label-free imaging, such as Raman microscopy [20][21][22] , focused ion beam scanning electron microscopy (called FIB-SEM) 23,24 and optical diffraction tomography combined with holographic microscopy (holotomographic microscopy), have been developed. Notably, holotomographic microscopy, which includes phase contrast microscopy, was developed to combine holographic and tomographic techniques based on measuring the refractive index (RI) of a sample [25][26][27][28] . The best characteristic of this technique is that chemical or physical fixation is not necessary and holography can reproduce original objects with authenticity to provide three-dimensional (3D) structural information.…”

Impact of sample preparation on holographic imaging of leukocytes

“…Measurements were performed with a transmission off-axis holographic microscope provided by Ovizio Imaging Systems, Belgium, which has been described before. 14,16 It is a double-differential DHM and is equipped with a 40× NA 0.55 Nikon CFI LWD 40× Cremove objective, a 528-nm Oslon PowerStar SLED (Osram), and a PointGrey Grashopper GS3U332S4 camera (105 fps, exposure time 5 μs) for high-throughput imaging (Fig. 4, Appendix A).…”