Method for electron microscopic studies of circulating human leukocytes and observations on their fine structure

Watanabe, Itaru; Donahue, Sheila A.; Hoggatt, Norma

doi:10.1016/s0022-5320(67)80106-0

Cited by 88 publications

(34 citation statements)

References 9 publications

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“…These findings are in good accordance with the excellent descriptions by ANDERSON (1966) andWATANABE, DONAHUE andHOGGART (1967).…”

Section: Results and Commentssupporting

confidence: 93%

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A Simple Micromethod of Preparing Peripheral Leukocytes and Platelets for Electron Microscopy

Hattori

1970

Arch. Histol. Jap., Arch. Histol. Jpn

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“…These findings are in good accordance with the excellent descriptions by ANDERSON (1966) andWATANABE, DONAHUE andHOGGART (1967).…”

Section: Results and Commentssupporting

confidence: 93%

“…The buffy coat thus hardened was taken out as a disc from the tube and cut into small pieces to be treated thereafter as usual tissue blocks. This and similar methods (MIURA et al, 1966;WATANABE, DONAHUE and HOGGART, 1967;WHITE, 1967) have been employed by an increasing number of investigators because of their simplicity and consistent results.…”

mentioning

confidence: 99%

A Simple Micromethod of Preparing Peripheral Leukocytes and Platelets for Electron Microscopy

Hattori

1970

Arch. Histol. Jap., Arch. Histol. Jpn

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“…For in vitro studies involving normal peripheral blood eosinophils, eosinophils were isolated from the peripheral blood of normal and atopic donors by using immunomagnetic beads coupled to an anti-CD16 antibody as described (25). The (26). Grey to silver sections were cut with a Reichert-Jung Ultracut microtome (Leicha, Deerfield, IL), stained with uranyl and lead salts, and examined in a JEOL 100 CX microscope (JEOL) operating at 80 kV.…”

mentioning

confidence: 99%

Constitutive production of granulocyte/macrophage colony-stimulating factor by hypodense mononuclear eosinophils developed in vitro from hybrid eosinophil/basophil granulocytes.

Boyce¹,

Friend²,

Gurish³

et al. 1996

Proc. Natl. Acad. Sci. U.S.A.

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We recently described the development in vitro of cells with granules characteristic of eosinophils and basophils (hybrid granulocytes) from normal human cord blood mononuclear cells cultured for 14 days with recombinant human (rh) interleukin (IL)-3, rhIL-5, and a soluble basement membrane, Matrigel. Hybrid granulocytes constitutively produced granulocyte/macrophage colony-stimulating factor (GM-CSF) and rapidly developed into eosinophils after the exogenous cytokines and Matrigel were removed. To characterize the developmental progression of hybrid granulocytes, cells were maintained for an additional 14 days in medium containing rhIL-3, rhIL-5, and Matrigel. After 28 days, 73% ± 1% (mean + SEM; n = 6) of the nonadherent cells were mononuclear eosinophils, 13% + 3% were eosinophils with two or more nuclear lobes, 13% + 4% were hybrid granulocytes, and 0.2% ± 0.1% were basophils. More than 90%Yo of the mononuclear eosinophils were hypodense as determined by centrifugation through metrizamide gradients. After an additional 5 days of culture in medium without exogenous cytokines, 65% + 3% (n = 5) of the 28-day cells excluded trypan blue. In contrast, 2% ± 1% of freshly isolated peripheral blood eosinophils survived 5 days of culture without exogenous cytokines (n = 5). Fifty percent conditioned medium from in vitro derived 28-day mononuclear eosinophils and 14-day hybrid granulocytes maintained the survival of 60% + 7% and 77% + 7%, respectively, of freshly isolated peripheral blood eosinophils for 72 h, compared with 20%o + 8% survival in medium alone (n = 3). The eosinophil viabilitysustaining activity of 50%o mononuclear eosinophil-conditioned medium was neutralized with a GM-CSF antibody. A total of 88% of the 28-day cells exhibited immunochemical staining for GM-CSF. Thus, during eosinophilopoiesis, both hybrid eosinophil/basophil intermediates and immature mononuclear eosinophils exhibit autocrine regulation of viability due to constitutive production of GM-CSF.When centrifuged through discontinuous gradients of Percoll or metrizamide, peripheral blood eosinophils segregate into populations of normal (normodense) or relatively less (hypodense) centrifugation density (1). Whereas normodense eosinophils are the predominant physical phenotype found in the peripheral blood of healthy individuals, variable numbers of hypodense eosinophils are found in the blood of patients with eosinophil-associated diseases, such as the idiopathic hypereosinophilic syndrome (IHES) (2-4). This physical diversity is associated with functional heterogeneity (4-8). In comparison to normodense eosinophils, hypodense eosinophils exhibit transiently enhanced ex vivo survival attributable to exposure to eosinophilopoietic cytokines in vivo (4). Both normodenseThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact. and hypodense peripheral blood eosinophils maintain their viabi...

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“…Blood samples were collected in heparinized test tubes and kept at room temperature for 30 min, fol lowing which the buffy coat was removed and pro cessed according to Watanahe et al [17]. Aspirated bone marrow (1 case) was processed according to Ackerman and Clark [1].…”

Section: Methodsmentioning

confidence: 99%

Ultrastructure of Human Eosinophils Genetically Lacking Peroxidase

Presentey¹

1984

Acta Haematol

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Method for electron microscopic studies of circulating human leukocytes and observations on their fine structure

Cited by 88 publications

References 9 publications

A Simple Micromethod of Preparing Peripheral Leukocytes and Platelets for Electron Microscopy

A Simple Micromethod of Preparing Peripheral Leukocytes and Platelets for Electron Microscopy

Constitutive production of granulocyte/macrophage colony-stimulating factor by hypodense mononuclear eosinophils developed in vitro from hybrid eosinophil/basophil granulocytes.

Ultrastructure of Human Eosinophils Genetically Lacking Peroxidase

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