ABSTRACT
<p><b>OBJECTIVE</b>To investigate the Raman spectral characteristics of leukemia cells from 4 patients with acute promyelocytic leukemia (M) and 3 patients with acute monoblastic leukemia (M), establish a novel Raman label-free method to distinguish 2 kinds of acute myeloid leukemia cells so as to provide basis for clinical research.</p><p><b>METHODS</b>Leukemia cells were collected from bone marrow of above-mentioned patients. Raman spectra were acquired by Horiba Xplora Raman spectrometer and Raman spectra of 30-50 cells from each patient were recorded. The diagnostic model was established according to principle component analysis (PCA), discriminant function analysis (DFA) and cluster analysis, and the spectra of leukemia cells from 7 patients were analyzed and classified. Characteristics of Raman spectra were analyzed combining with ultrastructure of leukemia cells.</p><p><b>RESULTS</b>There were significant differences between Raman spectra of 2 kinds of leukemia cells. Compared with acute monoblastic leukemia cells, the spectra of acute promyelocytic leukemia cells showed stronger peaks in 622, 643, 757, 852, 1003, 1033, 1117, 1157, 1173, 1208, 1340, 1551, 1581 cm. The diagnostic models established by PCA-DFA and cluster analysis could successfully classify these Raman spectra of different samples with a high accuracy of 100% (233/233). The model was evaluated by "Leave-one-out" cross-validation and reached a high accuracy of 97% (226/233).</p><p><b>CONCLUSION</b>The level of macromolecules of Mcells is higher than that of M. The diagnostic models established by PCA-DFA can classify these Raman spectra of different cells with a high accuracy. Raman spectra shows consistent result with ultrastructure by TEM.</p>
ABSTRACT
<p><b>OBJECTIVE</b>To analyze and evaluate the application of spinning disk confocal microscopy and imaging analysis software in movement and phagocytosis of neutrophils.</p><p><b>METHODS</b>Neutrophils were isolated from bone marrow by centrifugation on discontinuous Percoll gradient, and then were stained with PE Gr-1 antibody and mixed with FITC-labeled Zymosan A bioparticles. Multichannel time-lapse videos were captured by using the spinning disk confocal microscopy. The result was analyzed by using volocity and ImageJ software, the parameters associated with movement and phagocytosis of neutrophils were analyzed, including morphological changes, cell tracking, pseudopod dynamics, binding and phagocytosis index.</p><p><b>RESULTS</b>Most neutrophils would be polarized in response to Zymosan particles during a short time. Binding and phagocytosis process occured in forty minutes.</p><p><b>CONCLUSION</b>A method of precisely quantifying the movement and phagocytosis of neutrophils using microscopic imaging and imaging analysis technique has been set up successfully. Using this method, biological activity and function of neutrophils can be evaluated visually and rapidly. The physiologically rapid response to Zymosan particles can be applied to the neutrophils function research in the future.</p>