Dataset Information

Improved methods for magnetic purification of malaria parasites and haemozoin.

ABSTRACT:

Background

Malaria parasites generate free haem upon catabolism of host haemoglobin during their intraerythrocytic growth cycle. In order to minimize oxidative toxicity of the ferric iron, the free haem molecules are polymerized into the biomineral beta-haematin (commonly referred to as haemozoin). Haemozoin crystals are paramagnetic, and this property can be exploited for the purification of late stage parasites as they contain larger haemozoin crystals than early stage parasites and uninfected cells. Commercially available magnets that were originally developed for the purpose of antibody-mediated cell purification are widely used for this purpose. As these methods are not necessarily optimized for parasite purification, the relationship between magnetic field strength and the quantity and quality of yield during parasite purification was explored.

Methods

Inexpensive rare-earth neodymium magnets with commercially available disposable columns were employed to explore the relationship between magnetic field strength and recovery of free haemozoin and infected erythrocytes (iRBCs).

Results

Yields of free haemozoin increased nearly linearly with increasing magnetic field strength to the strongest fields tested (8,500 Gauss). Stronger magnetic fields also improved the recovery of iRBCs with no detrimental effects on parasite viability. An in-house constructed magnetic stand, built for $75 in materials, produced superior results when compared with much more expensive commercial products.

Conclusions

Existing protocols for the magnetic purification of free haemozoin and iRBCs result in sub-optimal yields. Inexpensive high-strength neodymium magnets offer a better option, resulting in higher yields with no detrimental effects on parasite viability.

SUBMITTER: Kim CC

PROVIDER: S-EPMC2817699 | biostudies-literature | 2010 Jan

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Improved methods for magnetic purification of malaria parasites and haemozoin.

Kim Charles C CC Wilson Emily B EB DeRisi Joseph L JL

Malaria journal 20100114

<h4>Background</h4>Malaria parasites generate free haem upon catabolism of host haemoglobin during their intraerythrocytic growth cycle. In order to minimize oxidative toxicity of the ferric iron, the free haem molecules are polymerized into the biomineral beta-haematin (commonly referred to as haemozoin). Haemozoin crystals are paramagnetic, and this property can be exploited for the purification of late stage parasites as they contain larger haemozoin crystals than early stage parasites and un ...[more]

PMID: 20074366

Similar Datasets

Project description:BackgroundDespite intensive research, malaria remains a major health concern for non-immune residents and travelers in malaria-endemic regions. Efficient adjunctive therapies against life-threatening complications such as severe malarial anaemia, encephalopathy, placental malaria or respiratory problems are still lacking. Therefore, new insights into the pathogenesis of severe malaria are imperative. Haemozoin (Hz) or malaria pigment is produced during intra-erythrocytic parasite replication, released in the circulation after schizont rupture and accumulates inside multiple organs. Many in vitro and ex vivo immunomodulating effects are described for Hz but in vivo data are limited. This study aimed to improve methods for Hz quantification in tissues and to investigate the accumulation of Hz in different organs from mice infected with Plasmodium parasites with a varying degree of virulence.MethodsAn improved method for extraction of Hz from tissues was elaborated and coupled to an optimized, quantitative, microtiter plate-based luminescence assay with a high sensitivity. In addition, a technique for measuring Hz by semi-quantitative densitometry, applicable on transmitted light images, was developed. The methods were applied to measure Hz in various organs of C57BL/6 J mice infected with Plasmodium berghei ANKA, P. berghei NK65 or Plasmodium chabaudi AS. The used statistical methods were the Mann-Whitney U test and Pearsons correlation analysis.ResultsMost Hz was detected in livers and spleens, lower levels in lungs and kidneys, whereas sub-nanomolar amounts were observed in brains and hearts from infected mice, irrespectively of the parasite strain used. Furthermore, total Hz contents correlated with peripheral parasitaemia and were significantly higher in mice with a lethal P. berghei ANKA or P. berghei NK65-infection than in mice with a self-resolving P. chabaudi AS-infection, despite similar peripheral parasitaemia levels.ConclusionsThe developed techniques were useful to quantify Hz in different organs with a high reproducibility and sensitivity. An organ-specific Hz deposition pattern was found and was independent of the parasite strain used. Highest Hz levels were identified in mice infected with lethal parasite strains suggesting that Hz accumulation in tissues is associated with malaria-related mortality.

Project description:Background:Early and accurate diagnosis of malaria is critical to the success of malaria elimination. However, the current mainstay of malaria diagnosis in the field, such as light microscopy and rapid diagnostic tests (RDTs), have limitations due to low parasite density or mutation in diagnostic markers. Methods:We evaluated an inexpensive, robust, rapid, malaria diagnostic device, called Gazelle, that employs magneto-optical detection to identify haemozoin crystals (Hz) produced by all species of human malaria parasites in infected individuals. A beam of polarised light is passed through the lysed diluted blood sample under the influence of high (~.55T) and low magnetic fields. The difference in light transmission through the sample between the high and low magnetic fields indicates presence of Hz, suggesting possible malarial infection. A total of 300 febrile patients were screened at the malaria clinic of Indian Council of Medical Research-National Institute of Research in Tribal Health (ICMR-NIRTH), Jabalpur, India, from August 2018 to November 2018. Malaria diagnosis was done using four diagnostic methods: Gazelle, light microscopy, RDT, and malaria specific Polymerase Chain Reaction (PCR). Measures of diagnostic accuracy were compared. Findings:Out of 300 febrile patients enroled and tested for the presence of malaria parasites, 262 patient samples were included in the final analysis. The sensitivity and specificity of Gazelle was 98% and 97% in comparison to light microscopy, 82% and 99% to PCR and 78% and 99% to RDT, respectively. The results of the four diagnostic methods were comparable and statistically no significant differences in sensitivity or specificity was observed between these methods. Enhanced diagnostic accuracy of Gazelle in malaria patients with no prior history of malaria treatment was observed in this study. Interpretation:The diagnostic ability of Gazelle was comparable to light microscopy and better than RDTs even in low parasitemia and in presence of pfhrp2/3 deletion mutant parasites. Gazelle may be a novel valuable diagnostic tool in resource poor settings where (i) microscopy is not feasible and (ii) pfhrp2/3gene deleted parasite are present. Its speed, cost-efficiency, and alternative to lack of microscopists makes it an important adjunct in field settings. Funding:HemexDx, India.

Dataset Information

Improved methods for magnetic purification of malaria parasites and haemozoin.

Background

Methods

Results

Conclusions

Publications

Improved methods for magnetic purification of malaria parasites and haemozoin.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets