Detection of Plasmodia by Acridine Orange Stain

Richards, D F; Hunter, Dewitt T.; Janis, Burton

doi:10.1093/ajcp/51.2_ts.280

Cited by 7 publications

(4 citation statements)

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“…7 Strategies to diagnose malaria are based on detecting the parasite in the bloodstream. Besides conventional optical microscopy several diagnostic techniques have been developed for malaria detection including serological (dipstick) antigen detection, [8][9][10][11] fluorescence microscopy, [11][12][13][14][15] PCR-based assays, 11,16,17 flow cytometry, 11,18 NMR-based metabolic profiling 19 magneto-optic-based technology 20 and laser desorption mass spectrometry (LD-MS). 21,22 Hitherto, the potential of optical-based spectroscopic techniques as malaria diagnostics have not been fully exploited, although resonance Raman spectroscopy is showing potential due to the huge resonant enhancement of haemozoin bands.…”

Section: Introductionmentioning

confidence: 99%

Resonance Raman microscopy in combination with partial dark-field microscopy lights up a new path in malaria diagnostics

Wood

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Lasch

et al. 2009

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Our goal is to produce a rapid and accurate diagnostic tool for malaria using resonance Raman spectroscopy to detect small inclusions of haemozoin in Plasmodium falciparum infected red blood cells. In pursuit of this aim we serendipitously discovered a partial dark-field effect generated by our experimental setup, which helps identify in thick blood films potential parasites that are normally difficult to see with conventional bright-field microscopy. The haemozoin deposits 'light up' and these can be selectively targeted with the Raman microscope to confirm the presence or absence of haemozoin by the strong 1569 cm(-1) band, which is a marker for haemozoin. With newly developed imaging Raman microscopes incorporating ultra-sensitive rapid readout CCDs it is possible to obtain spectra with a good signal-to-noise ratio in 1 second. Moreover, images from a smear of potentially infected cells can be recorded and analysed with multivariate methods. The reconstructed images show what appear to be sub-micron-inclusions of haemozoin in some cells indicating that the technique has potential to identify low pigmented forms of the parasite including early trophozoite-stage infected cells. Further work is required to unambiguously confirm the presence of such forms through systematic staining but the results are indeed promising and may lead to the development of a new Raman-based malaria diagnostic.

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Section: Introductionmentioning

confidence: 99%

Resonance Raman microscopy in combination with partial dark-field microscopy lights up a new path in malaria diagnostics

Wood

Hermelink

Lasch

et al. 2009

Analyst

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“…Several diagnostic techniques have been employed for malaria detection, these include fluorescence microscopy, − polymerase chain reaction (PCR)-based assays, − serological (dipstick) antigen detection, ,− flow cytometry , and laser desorption mass spectrometry . Hitherto, the potential of vibrational spectroscopic techniques as malaria diagnostics have not been exploited, which is surprising given the important role the technique has played in understanding the molecular and electronic structure of β-hematin and Hz. ,,, In an early study by Slater et al Fourier transform infrared (FT-IR) spectroscopy was used to identify vibrational modes specifically associated with the propionate groups that link the heme groups together in the heme dimeric array .…”

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confidence: 99%

“…Several diagnostic techniques have been employed for malaria detection, these include fluorescence microscopy, [14][15][16][17][18] polymerase chain reaction (PCR)-based assays, [18][19][20] serological (dipstick) antigen detection, 18,[21][22][23] flow cytometry 18,24 and laser desorption mass spectrometry. 25 Hitherto, the potential of vibrational spectroscopic techniques as malaria diagnostics have not been exploited, which is surprising given the important role the technique has played in understanding the molecular and electronic structure of β-hematin and Hz.…”

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confidence: 99%

Discriminating the Intraerythrocytic Lifecycle Stages of the Malaria Parasite Using Synchrotron FT-IR Microspectroscopy and an Artificial Neural Network

et al. 2009

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Synchrotron Fourier transform infrared (FT-IR) spectra of fixed single erythrocytes infected with Plasmodium falciparum at different stages of the intraerythrocytic cycle are presented for the first time. Bands assigned to the hemozoin moiety at 1712, 1664, and 1209 cm(-1) are observed in FT-IR difference spectra between uninfected erythrocytes and infected trophozoites. These bands are also found to be important contributors in separating the trophozoite spectra from the uninfected cell spectra in principal components analysis. All stages of the intraerythrocytic lifecycle of the malarial parasite, including the ring and schizont stage, can be differentiated by visual inspection of the C-H stretching region (3100-2800 cm(-1)) and by using principal components analysis. Bands at 2922, 2852, and 1738 cm(-1) assigned to the nu(asym)(CH(2) acyl chain lipids), nu(sym)(CH(2) acyl chain lipids), and the ester carbonyl band, respectively, increase as the parasite matures from its early ring stage to the trophozoite and finally to the schizont stage. Training of an artificial neural network showed that excellent automated spectroscopic discrimination between P. falciparum-infected cells and the control cells is possible. FT-IR difference spectra indicate a change in the production of unsaturated fatty acids as the parasite matures. The ring stage spectrum shows bands associated with cis unsaturated fatty acids. The schizont stage spectrum displays no evidence of cis bands and suggests an increase in saturated fatty acids. These results demonstrate that different phases of the P. falciparum intraerthyrocytic life cycle are characterized by different lipid compositions giving rise to distinct spectral profiles in the C-H stretching region. This insight paves the way for an automated infrared-based technology capable of diagnosing malaria at all intraerythrocytic stages of the parasite's life cycle.

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“…Compared with the staining with DAPI alone, the coloration of cytoplasmic RNA with a subsequent PI-stain made the outline of the parasites clearer and increased the brightn ess of the field of vision. Furthermore , differential staining of nuclei with DAPI from cytoplasm with PI enabled one easier discrimination of the parasites than in singl e staining with any of the known fluorescent dyes such as DAPI (3 -5) , mithramycin (12), acridine orange (13,14), rhodamine (15,16), ethidium bromide (17) and auramine (18,19). It should be noted that double staining with DAPI -PI was inadequate to observe mitochondrial DNA except that in T .…”

Section: Discussionmentioning

confidence: 99%

Simple and Rapid Staining for Detection of Entamoeba Cysts and Other Protozoans With Fluorochromes

Kawamoto

Mizuno

Fujioka

et al. 1987

JJMSB

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Detection of Plasmodia by Acridine Orange Stain

Cited by 7 publications

References 0 publications

Resonance Raman microscopy in combination with partial dark-field microscopy lights up a new path in malaria diagnostics

Resonance Raman microscopy in combination with partial dark-field microscopy lights up a new path in malaria diagnostics

Discriminating the Intraerythrocytic Lifecycle Stages of the Malaria Parasite Using Synchrotron FT-IR Microspectroscopy and an Artificial Neural Network

Simple and Rapid Staining for Detection of Entamoeba Cysts and Other Protozoans With Fluorochromes

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