Microfluidics-Based Approaches to the Isolation of African Trypanosomes

Abstract

African trypanosomes are responsible for significant levels of disease in both humans and animals. The protozoan parasites are free-living flagellates, usually transmitted by arthropod vectors, including the tsetse fly. In the mammalian host they live in the bloodstream and, in the case of human-infectious species, later invade the central nervous system. Diagnosis of the disease requires the positive identification of parasites in the bloodstream. This can be particularly challenging where parasite numbers are low, as is often the case in peripheral blood. Enriching parasites from body fluids is an important part of the diagnostic pathway. As more is learned about the physicochemical properties of trypanosomes, this information can be exploited through use of different microfluidic-based approaches to isolate the parasites from blood or other fluids. Here, we discuss recent advances in the use of microfluidics to separate trypanosomes from blood and to isolate single trypanosomes for analyses including drug screening.

Lab-on-a-Chip Technologies for the Single Cell Level: Separation, Analysis, and Diagnostics

“…Some salient examples thereof are circulating tumor cells (CTC) of various types of cancer [1,[36][37][38], rare cells (e.g., sickle-cell variants of red blood cells) [39,40], parasites, like Plasmodium falciparum [1,7,10,11,[13][14][15][21][22][23]36,[41][42][43][44][45][46][47][48][49][50] and Trypanosoma spp. [8,44,[51][52][53][54][55][56][57] and even plant pathogens [26,58], as well as-after cells have been lysed-subcellular infection markers (e.g., DNA, RNA fragments) [10,11,22,43,[59][60][61][62]. Given the vast adaptability of microfluidics to any kind of single or multi-cellular assay [63], the ability to combine it with various light microscopy techniques…”

“…This movement is caused by the induction of an electric dipole moment on the cell or particle, and the force the electric field gradient exerts on this dipole moment. It is not necessary that a cell carries a net surface electric charge to be polarized [8]. The induced dipole is aligned along the gradient of the non-uniform electric field.…”

“…DEP can also be combined with other technologies for single cell diagnostic chips. One example is the combination of DEP with deterministic lateral displacement (DLD) as demonstrated by Jason Beech and others [6,8,[88][89][90]; another prime example is the combination with SAW (see Section 2.1.5. ), as used by Smith et al to extract viable mesenchymal stroma cells from human dental pulp [91].…”

“…This demonstrates that it is possible to completely separate different cell types from the same sample, simply based on their induced dipole moment. Since the induced dipole moment is specific for healthy cells (e.g., RBCs), infected cells (e.g., RBCs infected by Plasmodium falciparum), and pathogenic cells (e.g., trypanosomes) they can be separated in such an experimental setup [6,8,54,86,87] (see Figure 2). In many cases, the detection of a single infected cell or individual pathogen within a real-life sample can arguably be counted as a diagnosis of the respective infection (here: Malaria and Trypanosomiasis, respectively).…”

“…One very prominent aspect of microfluidic research is diagnostics on the single-cell or even molecular level. The significance of recent research towards microfluidics-based single cell diagnostic chips is apparent in health care, in our homes, and also very prominently in the fight against the COVID 19 pandemic: The diagnostic targets range from circulating tumor cells (CTC) [1][2][3][4][5][6], over parasites in blood [7][8][9][10][11][12][13][14][15], male fertility [16][17][18][19][20], molecular markers for infections [11,15,[21][22][23], cells of a specific stage in their life cycle [24,25], plant pathogens [26] and the SARS-CoV-2 proteome [27][28][29][30][31][32]. Depending on the exact target (either the entire cell or sub-cellular markers) there are different approaches to on-chip detection, each with their own underlying fundamentals and set of limitations.…”