An automated haematology analyzer XN-30 distinguishes developmental stages of falciparum malaria parasite cultured in vitro

Abstract

BackgroundThe automated haematology analyzer XN-30 (Sysmex, Kobe, Japan) easily and rapidly detects malarial parasites in clinical blood samples using flow cytometry. The XN-30 analyzer is able to distinguish each developmental stage by measuring DNA content and cell size. Thus, it was expected to be capable of quantifying the developmental stages of cultured falciparum parasite. To achieve this requirement, a modified algorithm was tested for its validity and reliability using in vitro cultured falciparum parasite.ResultsThe XN-30 analyzer automatically measured each developmental stage as well as total parasitaemia. Comparison of the parasitaemia obtained using the XN-30 analyzer equipped with the modified algorithm with that obtained using microscopy examination of Giemsa-stained smears revealed the greater sensitivity and reproducibility of the former. The XN-30 analyzer also detected free merozoites and purified gametocytes.ConclusionsThe XN-30 analyzer allows the precise recognition and enumeration of total and each developmental stages of cultured falciparum parasites, and permits the sensitive and reproducible calculation of parasitaemia. The results indicate the potential of the XN-30 analyzer for basic research on malarial biology, anti-malarial drug discovery, and evaluation of drug efficacy.Electronic supplementary materialThe online version of this article (10.1186/s12936-018-2208-6) contains supplementary material, which is available to authorized users.

A simple monochromatic flow cytometric assay for assessment of intraerythrocytic development of Plasmodium falciparum

“…The accurate detection and quantification of P. falciparum-infected erythrocytes using an automated Sysmex Haematology Analyzer XN-30 was recently reported [27,28]. This automatic machine employs a 405-nm laser to detect cells that need to be partially lysed to increase the permeability of florescent dye before cell analysis.…”

Application of the automated haematology analyzer XN-30 in an experimental rodent model of malaria

“…The XN-30 analyzer was previously reported to allow the automatic detection of iRBCs in human blood samples and in vitro cultures [ 7 , 8 ]. However, the default setting of the XN-30 analyzer is insufficient for the precise detection of iRBCs from mouse blood sample.…”

“…The minimum detection sensitivity of the XN-30 analyzer in the whole blood mode, which allows quantitative counting of infected RBCs is 18 iRBCs/μL [ 7 ]. In addition, a new algorithm for in vitro cultured parasites was developed, which distinguished the different developmental stages of in vitro cultured Plasmodium falciparum parasite [ 8 ]. Thus, it was inferred that the XN-30 analyzer may be suitable not only for easy and rapid parasitaemia measurements in human blood or in vitro parasite culture but could also be an important experimental tool for in vivo drug screening and development in rodent malarial models.…”

The malaria parasite Plasmodium falciparum in red blood cells selectively takes up serum proteins that affect host pathogenicity

“…Slides were observed under 1000× magnification using a model BX50 light microscope (Olympus, Tokyo, Japan). An automated haematology analyser XN-30 equipped with an algorithm for cultured P. falciparum parasites (prototype: software version 01-03, build 16) was used with dedicated reagents (CELLPACK DCL, SULFOLYSER, Lysercell M, and Fluorocell M) (Sysmex, Kobe, Japan) [29]. Approximately 100 µL of culture suspension was aliquoted into a standard 1.5 mL microtube (Eppendorf Tubes 3810X; Eppendorf, Hamburg, Germany), and loaded into the XN-30 analyser as described in the manufacturer's manual (Sysmex).…”

“…Approximately 100 µL of culture suspension was aliquoted into a standard 1.5 mL microtube (Eppendorf Tubes 3810X; Eppendorf, Hamburg, Germany), and loaded into the XN-30 analyser as described in the manufacturer's manual (Sysmex). Parasitaemia (total, MI-RBC%; ring-form, RNG-RBC%; trophozoite, TRPZ-RBC%; and schizont, SCHZ-RBC%) was automatically reported [29]. However, as the XN-30 analyser detects DNA content, parasitaemia was overestimated due to the presence of dead parasites.…”