Leukocyte lipid bodies: inflammation-related organelles are rapidly detected by wet scanning electron microscopy

Leukocyte lipid bodies are dynamic, functionally active organelles with central roles in inflammation. Here, we report that leukocyte lipid bodies are facilely detected by a versatile, potent technique, termed wet scanning electron microscopy (SEM), which combines the rapid preparation of light microscopy with the resolution of SEM. Using as leukocyte models resting and agonist-stimulated human eosinophils, cells that generate prominent numbers of lipid bodies in inflammatory conditions, we demonstrated that lipid bodies can be rapidly imaged as bright, highly contrasted structures under wet SEM and scored by computerized image processing. Critical advantages of this approach are that it permits cell observation in a fully hydrated system and facilitates lipid preservation. These attributes are especially important because lipid bodies are degraded during routine dehydration processes. Moreover, this technology is advantageous over lipophilic fluorescent probes because it allows sustained detection of lipid bodies in contrast to short-lived fluorescent labeling of these organelles. The value of wet SEM in enabling rapid and large-scale lipid body imaging and scoring within leukocytes is particularly important because lipid bodies are organelles underlying the heightened functions of inflammatory cells. Wet SEM technology provides new approaches and opportunities for delineations of lipid bodies in inflammatory diseases, including allergic inflammation. Leukocyte lipid bodies are dynamic, functionally active organelles with central roles in inflammation. Here, we report that leukocyte lipid bodies are facilely detected by a versatile, potent technique, termed wet scanning electron microscopy (SEM), which combines the rapid preparation of light microscopy with the resolution of SEM. Using as leukocyte models resting and agonist-stimulated human eosinophils, cells that generate prominent numbers of lipid bodies in inflammatory conditions, we demonstrated that lipid bodies can be rapidly imaged as bright, highly contrasted structures under wet SEM and scored by computerized image processing. Critical advantages of this approach are that it permits cell observation in a fully hydrated system and facilitates lipid preservation. These attributes are especially important because lipid bodies are degraded during routine dehydration processes. Moreover, this technology is advantageous over lipophilic fluorescent probes because it allows sustained detection of lipid bodies in contrast to short-lived fluorescent labeling of these organelles. The value of wet SEM in enabling rapid and large-scale lipid body imaging and scoring within leukocytes is particularly important because lipid bodies are organelles underlying the heightened functions of inflammatory cells. Wet SEM technology provides new approaches and opportunities for delineations of lipid bodies in inflammatory diseases, including allergic inflammation. Abbreviations BODIPY2-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-pentanoyl)-1-hexadecanoyl-sn-glycero-3-phosphate, diammonium saltBSEbackscattered electronSEMscanning electron microscopyTEMtransmission electron microscopy 2-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-pentanoyl)-1-hexadecanoyl-sn-glycero-3-phosphate, diammonium salt backscattered electron scanning electron microscopy transmission electron microscopy The organization of lipids bearing esterified arachidonic acid into distinct intracellular sites has particularly important roles in cells of the immune system (1Yaqoob P. Lipids and the immune response: from molecular mechanisms to clinical applications..Curr. Opin. Clin. Nutr. Metab. Care. 2003; 6: 133-150Crossref PubMed Scopus (69) Google Scholar). In leukocytes and other cells of the inflammatory response, arachidonyl lipids are often discretely organized within cytoplasmic lipid bodies and serve as precursors of inflammatory eicosanoid mediators (2Dvorak A.M. Dvorak H.F. Peters S.P. Shulman E.S. MacGlashan Jr., D.W. Pyne K. Harvey V.S. Galli S.J. Lichtenstein L.M. Lipid bodies: cytoplasmic organelles important to arachidonate metabolism in macrophages and mast cells..J. Immunol. 1983; 131: 2965-2976PubMed Google Scholar, 3Weller P.F. Bozza P.T. Yu W. Dvorak A.M. Cytoplasmic lipid bodies in eosinophils: central roles in eicosanoid generation..Int. Arch. Allergy Immunol. 1999; 118: 450-452Crossref PubMed Scopus (53) Google Scholar, 4Bozza P.T. Bandeira-Melo C. Mechanisms of leukocyte lipid body formation and function in inflammation..Mem. Inst. Oswaldo Cruz. 2005; 100 (Suppl. 1):: 113-120Crossref PubMed Scopus (37) Google Scholar, 5Weller P.F. Dvorak A.M. Lipid bodies: intracellular sites for eicosanoid formation..J. Allergy Clin. Immunol. 1994; 94: 1151-1156Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar). After in vitro inflammatory stimuli or during in vivo inflammatory diseases, including infectious diseases, the numbers and sizes of lipid bodies increase within inflammatory cells (e.g., neutrophils, eosinophils, and macrophages), and these organelles become predominant sites for the synthesis of eicosanoid inflammatory mediators (3Weller P.F. Bozza P.T. Yu W. Dvorak A.M. Cytoplasmic lipid bodies in eosinophils: central roles in eicosanoid generation..Int. Arch. Allergy Immunol. 1999; 118: 450-452Crossref PubMed Scopus (53) Google Scholar, 6D'Avila H. Melo R.C.N Parreira G.G. Werneck-Barroso E. Castro-Faria-Neto H.C. Bozza P.T. Mycobacterium bovis bacillus Calmette-Guerin induces TLR2-mediated formation of lipid bodies: intracellular domains for eicosanoid synthesis in vivo..J. Immunol. 2006; 176: 3087-3097Crossref PubMed Scopus (233) Google Scholar, 7Bandeira-Melo C. Phoofolo M. Weller P.F. Extranuclear lipid bodies, elicited by CCR3-mediated signaling pathways, are the sites of chemokine-enhanced leukotriene C4 production in eosinophils and basophils..J. Biol. Chem. 2001; 276: 22779-22787Abstract Full Text Full Text PDF PubMed Scopus (107) Google Scholar, 8Melo R.C.N D'Avila H. Fabrino D.L. Almeida P.E. Bozza P.T. Macrophage lipid body induction by Chagas disease in vivo: putative intracellular domains for eicosanoid formation during infection..Tissue Cell. 2003; 35: 59-67Crossref PubMed Scopus (90) Google Scholar). Lipid bodies within inflammatory cells contain all of the enzymes necessary for eicosanoid synthesis, including cyclooxygenases, 5-and 15-lipoxygenases, and leukotriene C4-synthase (3Weller P.F. Bozza P.T. Yu W. Dvorak A.M. Cytoplasmic lipid bodies in eosinophils: central roles in eicosanoid generation..Int. Arch. Allergy Immunol. 1999; 118: 450-452Crossref PubMed Scopus (53) Google Scholar, 4Bozza P.T. Bandeira-Melo C. Mechanisms of leukocyte lipid body formation and function in inflammation..Mem. Inst. Oswaldo Cruz. 2005; 100 (Suppl. 1):: 113-120Crossref PubMed Scopus (37) Google Scholar, 6D'Avila H. Melo R.C.N Parreira G.G. Werneck-Barroso E. Castro-Faria-Neto H.C. Bozza P.T. Mycobacterium bovis bacillus Calmette-Guerin induces TLR2-mediated formation of lipid bodies: intracellular domains for eicosanoid synthesis in vivo..J. Immunol. 2006; 176: 3087-3097Crossref PubMed Scopus (233) Google Scholar, 9Dvorak A.M. Cyclooxygenase, a key enzyme family for production of prostaglandins, is present in human mast cell lipid bodies..Chem. Immunol. Allergy. 2005; 85: 68-71Crossref PubMed Scopus (2) Google Scholar). Both lipid body formation and the compartmentalization of enzymes within lipid bodies are highly regulated cellular events involved in the heightened capacity of leukocytes to generate eicosanoids. Lipid bodies, therefore, are dynamic, functionally active organelles that both contribute to the genesis of inflammatory and immune responses and act as targets for the control of inflammatory diseases. Although lipid bodies have central roles in inflammation and are considered structural markers of inflammatory cells in a range of diseases (6D'Avila H. Melo R.C.N Parreira G.G. Werneck-Barroso E. Castro-Faria-Neto H.C. Bozza P.T. Mycobacterium bovis bacillus Calmette-Guerin induces TLR2-mediated formation of lipid bodies: intracellular domains for eicosanoid synthesis in vivo..J. Immunol. 2006; 176: 3087-3097Crossref PubMed Scopus (233) Google Scholar, 10Melo R.C.N Fabrino D.L. Dias F.F. Parreira G.G. Lipid bodies: structural markers of inflammatory macrophages in innate immunity..Inflamm. Res. 2006; 55: 342-348Crossref PubMed Scopus (58) Google Scholar, 11Bozza P.T. Yu W. Penrose J.F. Morgan E.S. Dvorak A.M. Weller P.F. Eosinophil lipid bodies: specific, inducible intracellular sites for enhanced eicosanoid formation..J. Exp. Med. 1997; 186: 909-920Crossref PubMed Scopus (181) Google Scholar), their identification has methodological limitations because lipid bodies dissipate upon drying or dissolve upon fixation and staining with alcohol-based reagents (12Fukumoto S. Fujimoto T. Deformation of lipid droplets in fixed samples..Histochem. Cell Biol. 2002; 118: 423-428Crossref PubMed Scopus (144) Google Scholar, 13DiDonato D. Brasaemle D.L. Fixation methods for the study of lipid droplets by immunofluorescence microscopy..J. Histochem. Cytochem. 2003; 51: 773-780Crossref PubMed Scopus (109) Google Scholar, 14Ohsaki Y. Maeda T. Fujimoto T. Fixation and permeabilization protocol is critical for the immunolabeling of lipid droplet proteins..Histochem. Cell Biol. 2005; 124: 445-452Crossref PubMed Scopus (59) Google Scholar). For example, May-Grunwald-Giemsa staining causes the dissolution of lipid bodies (15Pacheco P. Bozza F.A. Gomes R.N. Bozza M. Weller P.F. Castro-Faria-Neto H.C. Bozza P.T. Lipopolysaccharide-induced leukocyte lipid body formation in vivo: innate immunity elicited intracellular loci involved in eicosanoid metabolism..J. Immunol. 2002; 169: 6498-6506Crossref PubMed Scopus (118) Google Scholar). Usually, these organelles are identified by light microscopy using lipophilic fluorescent probes (12Fukumoto S. Fujimoto T. Deformation of lipid droplets in fixed samples..Histochem. Cell Biol. 2002; 118: 423-428Crossref PubMed Scopus (144) Google Scholar, 13DiDonato D. Brasaemle D.L. Fixation methods for the study of lipid droplets by immunofluorescence microscopy..J. Histochem. Cytochem. 2003; 51: 773-780Crossref PubMed Scopus (109) Google Scholar, 16Fowler S.D. Greenspan P. Application of Nile Red, a fluorescent hydrophobic probe, for the detection of neutral lipid deposits in tissue sections: comparison with Oil Red O..J. Histochem. Cytochem. 1985; 33: 833-836Crossref PubMed Scopus (352) Google Scholar) or staining with osmium (17Bozza P.T. Payne J.L. Goulet J.L. Weller P.F. Mechanisms of platelet-activating factor-induced lipid body formation: requisite roles for 5-lipoxygenase and de novo protein synthesis in the compartmentalization of neutrophil lipids..J. Exp. Med. 1996; 183: 1515-1525Crossref PubMed Scopus (77) Google Scholar). At the ultrastructural level, lipid bodies can be observed by transmission electron microscopy (TEM) without any additional labeling, by which they appear as distinct organelles lacking delimiting classical trilaminar membranes (2Dvorak A.M. Dvorak H.F. Peters S.P. Shulman E.S. MacGlashan Jr., D.W. Pyne K. Harvey V.S. Galli S.J. Lichtenstein L.M. Lipid bodies: cytoplasmic organelles important to arachidonate metabolism in macrophages and mast cells..J. Immunol. 1983; 131: 2965-2976PubMed Google Scholar, 10Melo R.C.N Fabrino D.L. Dias F.F. Parreira G.G. Lipid bodies: structural markers of inflammatory macrophages in innate immunity..Inflamm. Res. 2006; 55: 342-348Crossref PubMed Scopus (58) Google Scholar, 18Dvorak A.M. Morgan E.S. Tzizik D.M. Weller P.F. Prostaglandin endoperoxide synthase (cyclooxygenase): ultrastructural localization to nonmembrane-bound cytoplasmic lipid bodies in human eosinophils and 3T3 fibroblasts..Int. Arch. Allergy Immunol. 1994; 105: 245-250Crossref PubMed Scopus (26) Google Scholar). However, TEM preparations are time-consuming and require costly processing, such as embedding and thin sectioning. Here, we report that leukocyte lipid bodies can be facilely detected by a wet scanning electron microscopy (SEM) technique, named WETSEM™, which enables the imaging of hydrated samples and combines the rapidity of preparation of light microscopy with the resolution of electron microscopy (19Thiberge S. Nechushtan A. Sprinzak D. Gileadi O. Behar V. Zik O. Chowers Y. Michaeli S. Schlessinger J. Moses E. Scanning electron microscopy of cells and tissues under fully hydrated conditions..Proc. Natl. Acad. Sci. USA. 2004; 101: 3346-3351Crossref PubMed Scopus (192) Google Scholar). Moreover, we have compared lipid body imaging by wet SEM with different cell preparations and staining methods for light microscopy. Using resting and agonist-stimulated human eosinophils, cells that can contain prominent numbers of lipid bodies (3Weller P.F. Bozza P.T. Yu W. Dvorak A.M. Cytoplasmic lipid bodies in eosinophils: central roles in eicosanoid generation..Int. Arch. Allergy Immunol. 1999; 118: 450-452Crossref PubMed Scopus (53) Google Scholar), as leukocyte models, we show that the wet SEM technology provides new approaches that facilitate the study of lipid bodies in inflammation. Paraformaldehyde (16%; electron microscopy grade), glutaraldehyde (25%; electron microscopy grade), uranyl acetate, and OsO4 were purchased from Electron Microscopy Sciences (Fort Washington, PA). Other chemicals, unless indicated, were obtained from Sigma (St. Louis, MO). Granulocytes were isolated from the blood of healthy donors as described (20Bandeira-Melo C. Gillard G. Ghiran I. Weller P.F. EliCell: a gel-phase dual antibody capture and detection assay to measure cytokine release from eosinophils..J. Immunol. Methods. 2000; 244: 105-115Crossref PubMed Scopus (30) Google Scholar), and eosinophils were purified by negative selection using human eosinophil enrichment cocktail (StemSep™; StemCell Technologies, Vancouver, Canada) and the magnetic activated cell sorter (MACS) bead procedure (Miltenyi Biotec, Auburn, CA). Sample procurement and experiments were approved by the Committee on Clinical Investigation, and informed consent was obtained from all subjects. Eosinophil purity was >99%. For different approaches, eosinophils (106 cells/ml) were stimulated with recombinant human eotaxin (100 ng/ml; R&D Systems, Minneapolis, MN) in RPMI-1640 medium plus 0.1% ovalbumin or medium alone at 37°C for 1 h (21Melo R.C.N Spencer L.A. Perez S.A.C Dvorak A.M. Weller P.F. Intragranular vesiculotubular compartments are involved in piecemeal degranulation by activated human eosinophils..Traffic. 2005; 6: 866-879Crossref PubMed Scopus (81) Google Scholar). Cell viability after stimulation was >95%, as determined by ethidium bromide incorporation. A nonphysiological challenge with the calcium ionophore A23187 (0.5 μM) was also used in some experiments, as described (22Bandeira-Melo C. Perez S.A. Melo R.C.N Ghiran I. Weller P.F. EliCell assay for the detection of released cytokines from eosinophils..J. Immunol. Methods. 2003; 276: 227-237Crossref PubMed Scopus (16) Google Scholar). Alternatively, unstimulated eosinophils were studied in whole samples of white blood cells isolated from human blood. Briefly, blood (3 ml) was mixed in a 6% dextran solution (1 ml) containing 3% glucose and 0.9% NaCl plus 60 μl of 0.5 M EDTA. After 15 min of sedimentation, the leukocyte-rich plasma layer was recovered, mixed with PBS, and centrifuged. White blood cells were resuspended in PBS and prepared for wet SEM. Suspensions containing nonstimulated and stimulated leukocytes were immediately placed into individual capsules (QuantomiX, Ltd.) (15 μl of cell suspension/capsule) containing a thin, electron-transparent partition membrane. Leukocytes were allowed to adhere to the membrane for 15 min at room temperature. Within the microscope, this membrane isolates the content of the capsules from the vacuum and allows both the penetration of electrons and the collection of backscattered electrons (BSEs) while withstanding a pressure difference of one atmosphere (19Thiberge S. Nechushtan A. Sprinzak D. Gileadi O. Behar V. Zik O. Chowers Y. Michaeli S. Schlessinger J. Moses E. Scanning electron microscopy of cells and tissues under fully hydrated conditions..Proc. Natl. Acad. Sci. USA. 2004; 101: 3346-3351Crossref PubMed Scopus (192) Google Scholar, 23Barshack I. Kopolovic J. Chowers Y. Gileadi O. Vainshtein A. Zik O. Behar V. A novel method for “wet” 2004; PubMed Google Scholar, A. Vainshtein A. J. Y. Gileadi O. Behar V. Electron microscopy of wet a study in 2004; PubMed Scopus Google Scholar). After cells were fixed with a prepared of and 0.1% glutaraldehyde in M PBS for min and in the OsO4 staining was by the fixed samples in and for min in OsO4 in After the samples with the capsules were Lipid bodies from unstimulated and human eosinophils were imaged by light microscopy with different Cell preparation was in using a (17Bozza P.T. Payne J.L. Goulet J.L. Weller P.F. Mechanisms of platelet-activating factor-induced lipid body formation: requisite roles for 5-lipoxygenase and de novo protein synthesis in the compartmentalization of neutrophil lipids..J. Exp. Med. 1996; 183: 1515-1525Crossref PubMed Scopus (77) Google Scholar) or by a of eosinophils with a L.A. Melo R.C.N Perez S.A. Weller P.F. A dual antibody capture and detection method for of cytokine Biol. 2005; Google Scholar). OsO4 staining and lipid body were as described (17Bozza P.T. Payne J.L. Goulet J.L. Weller P.F. Mechanisms of platelet-activating factor-induced lipid body formation: requisite roles for 5-lipoxygenase and de novo protein synthesis in the compartmentalization of neutrophil lipids..J. Exp. Med. 1996; 183: 1515-1525Crossref PubMed Scopus (77) Google Scholar) in both and Briefly, while were fixed with in in M in OsO4 in M in with M with OsO4 (3 and and Lipid bodies were using a in cells. In some experiments, preparations were with the after OsO4 For imaging of lipid bodies, eosinophils in preparations were with a and with 1 fluorescent 2-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-pentanoyl)-1-hexadecanoyl-sn-glycero-3-phosphate, diammonium salt for 1 h at 37°C fixation or with Nile Red from a solution of 1 in for min at room after Fixation was with in for After cells were in with and on a Alternatively, were after were with or microscopy at capsules were placed on the of the scanning electron to be in the wet After the imaging was to the of the microscope, with to some that from SEM imaging imaging was using a the electron be at 15 and is at and to for the the is a the be to The and different The in this were obtained on a or on a The scanning electron with a of or of of and of at were for lipid body using were compared by lipid bodies as as their within eosinophils, isolated human eosinophils were stimulated with eotaxin or medium alone for 1 h and immediately prepared for imaging with the wet SEM or by light microscopy. was because this is a potent eosinophil Mechanisms of Allergy Clin. Immunol. 2000; 105: Full Text Full Text PDF PubMed Scopus Google Scholar). activated by this or other stimuli in during inflammatory in with R.C.N Spencer L.A. Perez S.A.C Ghiran I. Dvorak A.M. Weller P.F. eosinophils distinct 2005; 6: PubMed Scopus Google Scholar), including the induction of new lipid body formation C. Phoofolo M. Weller P.F. Extranuclear lipid bodies, elicited by CCR3-mediated signaling pathways, are the sites of chemokine-enhanced leukotriene C4 production in eosinophils and basophils..J. Biol. Chem. 2001; 276: 22779-22787Abstract Full Text Full Text PDF PubMed Scopus (107) Google Scholar). light lipid bodies as or fluorescent organelles in cell preparations with osmium or fluorescent lipid probes In cells in compared with cells from preparations Moreover, a of activated human eosinophils R.C.N Weller P.F. Dvorak A.M. human Arch. Allergy Immunol. 2005; PubMed Scopus Google Scholar), be observed the cells were in Although lipid bodies were by osmium and this staining a in the eosinophil were In eosinophils prepared with Nile Red or lipid bodies were observed as or the was to of these organelles. Moreover, detection of lipid bodies with both fluorescent was in wet at the of the staining procedure show a of lipid were also These have from cellular by or cells of the as observed in other approaches using fluorescent L.A. Melo R.C.N Perez S.A. Weller P.F. A dual antibody capture and detection method for of cytokine Biol. 2005; Google Scholar). of lipid bodies with osmium of these organelles in cells lipid in unstimulated cells compared with lipid in eosinophils In both unstimulated and stimulated eosinophils, lipid bodies were by wet SEM as highly organelles Lipid bodies as white structures as observed for lipid imaging of wet samples using (19Thiberge S. Nechushtan A. Sprinzak D. Gileadi O. Behar V. Zik O. Chowers Y. Michaeli S. Schlessinger J. Moses E. Scanning electron microscopy of cells and tissues under fully hydrated conditions..Proc. Natl. Acad. Sci. USA. 2004; 101: 3346-3351Crossref PubMed Scopus (192) Google Scholar). In lipid bodies were distinct from which be as structures compared with by microscopy image scored lipid body numbers and a increase of in eosinophils compared with unstimulated cells lipid in unstimulated cells compared with lipid in eosinophils as also in this by light body imaging and within a human blood lipid bodies are observed in with within the In lipid bodies were scored by computerized image processing. were from a whole of white blood cells by wet SEM. was using image of the resolution of SEM compared with light a range of lipid body sizes was with increase in the of these organelles in activated cells The wet SEM also the cell and was the of lipid bodies to or in with the In contrast to which is on electron detection and to the cell the wet SEM enabling the of In this we have demonstrated that wet SEM technology is particularly for the rapid and of lipid bodies within human These organelles are sites of eicosanoid formation in leukocytes and have because of their with inflammatory diseases P.T. Bandeira-Melo C. Mechanisms of leukocyte lipid body formation and function in inflammation..Mem. Inst. Oswaldo Cruz. 2005; 100 (Suppl. 1):: 113-120Crossref PubMed Scopus (37) Google Scholar, P.F. Dvorak A.M. Lipid bodies: intracellular sites of eicosanoid In and Google Scholar, M. Cell of 2006; PubMed Scopus (81) Google Scholar). The highly contrasted lipid body imaging the wet SEM described for lipid body approach combines the advantages of light microscopy rapid the and the resolution of electron it be in that the resolution is compared with In because the wet SEM approach facilitates the of lipid this technology is advantageous over fluorescent lipid probes by detection in contrast to short-lived In lipid body can be a for routine of these organelles by especially is critical of wet SEM is that it permits cell observation in a fully hydrated is particularly important for lipid body because these organelles are highly to dehydration routine staining methods are for light microscopy (15Pacheco P. Bozza F.A. Gomes R.N. Bozza M. Weller P.F. Castro-Faria-Neto H.C. Bozza P.T. Lipopolysaccharide-induced leukocyte lipid body formation in vivo: innate immunity elicited intracellular loci involved in eicosanoid metabolism..J. Immunol. 2002; 169: 6498-6506Crossref PubMed Scopus (118) Google Scholar). Cell also from the wet SEM. we compared different of cell preparation and wet of leukocyte was in cells hydrated within the capsules or in the compared with preparations Wet SEM has to the study of different cells and In to lipid the is also for for rapid A. Vainshtein A. J. Y. Gileadi O. Behar V. Electron microscopy of wet a study in 2004; PubMed Scopus Google Scholar, I. S. Behar V. Vainshtein A. Zik O. E. M. Kopolovic J. D. Wet a novel method for rapid of 2004; PubMed Scopus Google Scholar) and imaging (19Thiberge S. Nechushtan A. Sprinzak D. Gileadi O. Behar V. Zik O. Chowers Y. Michaeli S. Schlessinger J. Moses E. Scanning electron microscopy of cells and tissues under fully hydrated conditions..Proc. Natl. Acad. Sci. USA. 2004; 101: 3346-3351Crossref PubMed Scopus (192) Google Scholar, A. M. D. A. I. E. A. Behar V. A new method of wet scanning electron microscopy for the of in of 2006; PubMed Scopus Google Scholar). lipid bodies, the of wet SEM is for rapid and large-scale lipid body imaging and scoring within leukocytes and other cells under different be a of leukocyte (15 is for the membrane enables the rapid of leukocytes (15 leukocytes are rapidly fixed and the capsules and can be imaged immediately after these capsules containing the samples are and can be at for because lipid bodies as structures under wet computerized image can the rapid of a of cells. light lipid body is after osmium Moreover, this staining in light microscopy especially in cells in which the is with and it lipid body in with lipid the wet SEM approach has the to be to lipid body in a range of cells. lipid bodies are present in cells and have other important with membrane and other regulated P. Y. Y. M. cell lipid droplets appear to be organelles involved in membrane Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, H. S. M. D. of and in lipid Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar, D. T. Y. J. V.S. Y. C. and lipid Cell Biol. 2006; PubMed Scopus Google Scholar). Wet SEM provides a imaging that can be used for lipid with for the study of leukocyte lipid bodies in inflammation. The of and Ghiran for and Behar (QuantomiX, Ltd.) for critical of the was by of and was by the de

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