Slide Veer Labs creative constructions for health malaria by mikeholy

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									                                       Detecting Birefringence of Hemozoin to Diagnose Malaria

                                                 Sheel M. Shah,1 Andrew M. Davis,1 Naman K. Shah,2 Steven R. Meshnick 2,3
                                             1 Department   s of Physics, 2 Epidemiology, 3 Microbiology & Immunology, University of North Carolina, Chapel Hill, NC, USA




                       ABSTRACT                                                                             RESULTS                                                              CONCLUSIONS
• Malaria infects roughly half a billion people every year,                                                                                                   • Sensitivity of signal and ease of detection is greatly
resulting in over a million deaths annually.                                                                                                                  increased with birefringence imaging

• As a host of inexpensive drug therapies exist to treat malaria,                                                                                             •Based on the increased signal sensitivity of using
the key to survival is early diagnosis and treatment 7.                                                                                                       birefringent hemozoin6, images produced via hemozoin
• Early diagnosis might be possible using the birefringence                                                                                                   birefringence could enable an automated, high-throughput,
property of hemozoin, a metabolic byproduct of the parasite.                                                                                                  low cost diagnosis device.

• We used a collimated polarized beam incident on an infected                                                                                                 • Diagnosis can be automated by translating a thick smear
sample to detect hemozoin with a polarizer completely out of                                                                                                  sample using stepper motors and imaging random fields.
phase with the polarization of the beam before the CCD.                Figure 1: Birefringent hemozoin superimposed on the same field as seen with ordinary
                                                                                  transmitted light.
• The increased sensitivity of the signal and ease of detection                                                                                                            FUTURE DIRECTIONS
lends itself to automated algorithms for diagnosis.                        Figure 2 shows an image acquired over two channels. The
                                                                                                                                                              • Develop a full working prototype with images being
                                                                           red channel carried the transmitted light image and the green
                                                                                                                                                              exported to image analysis software loaded onto blackfin tm
                                                                           channel carried the polarized light image. The images were
                                                                                                                                                              (Analog Devices) Processor.
                                                                           superimposed using Adobe Photoshoptm .The hemozoin
                    BACKGROUND                                             crystals can be clearly seen as the green artifacts on the red
                                                                                                                                                              •Determine correlation of the parasitemia level to hemozoin
                                                                           background. Machine detection of the presence of the
 • Current Diagnosis Techniques – Rapid Diagnostic Tests                                                                                                      density in the blood smear
                                                                           parasite in blood samples is greatly improved using
 (RDT) utilize antigen detection, but have variable sensitivity,
                                                                           birefringence detection. We successfully imaged hemozoin
 low shelf life, and high cost. Microscopy is limited by the                                                                                                  •Test the prototype with infected whole blood in the field.
                                                                           in samples with parasitemia as low as 0.1%.
 need for trained personnel to perform the test.
                                                                                                                                                                         ACKNOWLEDGEMENTS
 • There is a need to develop an automated, high-throughput,
 accurate, and inexpensive device that will diagnose malaria                                                                                                  Annie Purfield and Jaina Patel for slide preparation and
 rapidly and efficiently.                                                                                                                                     maintaining parasite cultures. Dr. Micheal Chua and
                                                                                                                                                              Dr. Bob Bagnell for guidance in the microscopy
                        METHODS                                                                                                                               facilities. Dr. Robert Dennis of TESLa Laboratories for
                                                                                                                                                              providing engineer space and materials. Dr. Jungsang
 • P.falciparum (3D7) parasites from in vitro culture were                                                                                                    Kim for general guidance with the project and feedback
 reconstituted with packed RBC’s. 30 μL was dried on a slide                                                                                                  of methods
 as a thick smear and methanol fixed for 30 seconds.

 •Images were taken using a Nikon TE2000 Inverted
 microscope with a 20x objective and a 10x eye piece.                                                                                                                             REFERENCES
 Images were acquired using the DVC-1412 monochrome
 CCD camera.
                                                                                                                                                              1.   Scott C, van Zyl D, Ho E, Meyersfeld D, Ruvio L, Mendelow B,
 • The collimated beam was polarized uniaxially and a                                                                                                              Coetzer T, 2003. Automated Detection of Malaria-Associated
 polarizer was inserted before the CCD in an orientation                                                                                                           Intraleucocytic Haemozoin by Cell-Dyn CD4000 Depolarization
 completely out of phase with the polarization of the beam.                                                                                                        Analysis. Clin Lab Haematol 25.2: 77--86.
                                                                                                                                                              2.   de Langen A, van Dillen J, de Witte P, Muchetto S, Nagelkerke
                                                                                                   Figure 2: Initial Device Prototype                              N, Kager P, 2003. Detection of Malaria Pigment: Feasibility for
 • Image analysis was performed using software based on                                                                                                            Malaria Diagnosing in an Area with Seasonal Malaria in
 ImageJ (NIH, Bethesda). Noise threshold levels were                           Figure 2 shows the initial device prototype. The                                    Northern Namibia. Trop Med Int Health 11.6: 809--816.
 calibrated using the images from the imaged P. falciparum                     detection of these crystals was easily automated using                         3.   Noland G, Briones N, Sullivan D Jr, 2003. The Shape and Size
 negative samples. The area used for particle counting was                     few components:                                                                     of Hemozoin Crystals Distinguishes Diverse Plasmodium
 based on the size of hemozoin5.                                               • Stepper motors and flexible drive shafts to provide                               Species. Mol Biochem Parasitol 130.2: 91--99.
                                                                               sample translation                                                             4.   Lawrence C, 1999. Remembered: Malaria Haemozoin in
                                                                               •An inexpensive microscope for imaging                                              Leucocytes. Lancet 353.9167: 1852.
                                                                               • Standard polarizing film.                                                    5.   Ross N, Pritchard C, Rubin D, Dusé A, 2006. Automated
                                                                                                                                                                   Image Processing Method for the Diagnosis and Classification
                                                                                                                                                                   of Malaria on Thin Blood Smears. Med Biol Eng Compu 44:
                                                                                                                                                                   472--436.

								
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