Flow cytometric discrimination of seven lineage markers by using two fluorochromes

Boin, Francesco; Torchia, Maria Letizia Giardino; Borrello, Ivan; Noonan, Kimberly; Neil, Matthew S.; Soloski, Mark J.; Cimbro, Raffaello

doi:10.1371/journal.pone.0188916

Cited by 4 publications

(3 citation statements)

References 19 publications

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“…Analysis of cell profiles was conducted on a BD LSRFortessa (BD Biosciences) and the data was analyzed using FlowJo v10 (FlowJo LLC, Ashland, OR). Cell populations were determined using a gating strategy that eliminates dead cells, debris and doublets to identify T cells (CD3 + ), B cells (CD19 + ) 61 , NK cells (CD3 − , CD19 − , NK 1.1 + ) 62 , dendritic cells (CD3 − , CD19 − , NK1.1 − , CD11c + , CD11b + ), neutrophils (CD3 − , CD19 − , NK1.1 − , Ly6G + , CD11b + ,CD11c − ), eosinophils (CD3 − , CD19 − , NK1.1 − , CD11c − , Ly6G − , CD11b + , Ly6C + , SSC Hi ), monocytes (CD3 − , CD19 − , NK1.1 − , CD11c − , Ly6G − , CD11b + , SSC lo , Ly6C lo , F4/80 lo ) and macrophages (CD3 − , CD19 − , NK1.1 − , CD11c − , Ly6G − , CD11b + , SSC lo , Ly6C lo , F4/80 hi ) 63 (Supplementary Fig. 1 ).…”

Section: Methodsmentioning

confidence: 99%

Onchocerca volvulus bivalent subunit vaccine induces protective immunity in genetically diverse collaborative cross recombinant inbred intercross mice

et al. 2021

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This study tests the hypothesis that an Onchocerca volvulus vaccine, consisting of two recombinant antigens (Ov-103 and Ov-RAL-2) formulated with the combination-adjuvant Advax-2, can induce protective immunity in genetically diverse Collaborative Cross recombinant inbred intercross mice (CC-RIX). CC-RIX lines were immunized with the O. volvulus vaccine and challenged with third-stage larvae. Equal and significant reductions in parasite survival were observed in 7 of 8 CC-RIX lines. Innate protective immunity was seen in the single CC-RIX line that did not demonstrate protective adaptive immunity. Analysis of a wide array of immune factors showed that each line of mice have a unique set of immune responses to vaccination and challenge suggesting that the vaccine is polyfunctional, inducing different equally-protective sets of immune responses based on the genetic background of the immunized host. Vaccine efficacy in genetically diverse mice suggests that it will also be effective in genetically complex human populations.

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Section: Methodsmentioning

confidence: 99%

Onchocerca volvulus bivalent subunit vaccine induces protective immunity in genetically diverse collaborative cross recombinant inbred intercross mice

et al. 2021

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“…Then, the cells were washed with PBS and samples were analyzed by flow cytometry using FACSCanto II flow cytometer equipped with the FACSDiva software (BD Biosciences, San Jose, USA), from 50,000 events for each sample. The monocyte and lymphocyte population were distinguished based on FSC/SSC properties, and gating strategy protocol was performed as described in the literature [ 25 ], and illustrated in Additional file 1A .…”

Section: Methodsmentioning

confidence: 99%

Role of crotoxin in coagulation: novel insights into anticoagulant mechanisms and impairment of inflammation-induced coagulation

Gimenez

Cezarette

Bomfim

et al. 2020

J. Venom. Anim. Toxins incl. Trop. Dis

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Background: Snake venom phospholipases A 2 (svPLA 2) are biologically active toxins, capable of triggering and modulating a wide range of biological functions. Among the svPLA 2 s, crotoxin (CTX) has been in the spotlight of bioprospecting research due to its role in modulating immune response and hemostasis. In the present study, novel anticoagulant mechanisms of CTX, and the modulation of inflammation-induced coagulation were investigated. Methods: CTX anticoagulant activity was evaluated using platelet poor plasma (PPP) and whole blood (WB), and also using isolated coagulation factors and complexes. The toxin modulation of procoagulant and pro-inflammatory effects was evaluated using the expression of tissue factor (TF) and cytokines in lipopolysaccharide (LPS)-treated peripheral blood mononuclear cells (PBMC) and in WB. Results: The results showed that CTX impaired clot formation in both PPP and WB, and was responsible for the inhibition of both intrinsic (TF/factor VIIa) and extrinsic (factor IXa/factor VIIIa) tenase complexes, but not for factor Xa and thrombin alone. In addition, the PLA 2 mitigated the prothrombinase complex by modulating the coagulation phospholipid role in the complex. In regards to the inflammation-coagulation cross talk, the toxin was capable of reducing the production of the pro-inflammatory cytokines IL-1β, IL-6 and TNF-α, and was followed by decreased levels of TF and procoagulant activity from LPS-treated PBMC either isolated or in WB. Conclusion: The results obtained in the present study recognize the toxin as a novel medicinal candidate to be applied in inflammatory diseases with coagulation disorders.

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“…Regarding lymphocytes, only CD3 + cells (T cells) were analysed. Among them, CD8 + or γδ + cells were selected based on the presence of these markers, and CD3 + CD8 − γδ − cells were assigned as CD4 + lymphocytes [25].…”

Section: Methodsmentioning

confidence: 99%

CX3CL1–CX3CR1 Axis: A New Player in Coeliac Disease Pathogenesis

et al. 2019

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Background: The CX3CL1–CX3CR1 axis has been related to numerous diseases. The aim of our study was to investigate its involvement in coeliac disease (CD) pathogenesis, particularly in the early phase of the disease. Methods: We collected peripheral blood from CD patients and controls, enrolled in a 3-day gluten challenge, to study soluble CX3CL1, I-TAC and MIG by Luminex, CX3CL1 and CX3CR1 gene expression by qPCR, and CX3CR1 protein expression in monocytes and CD8+, CD4+ and γδ+ T cells, by flow cytometry. We also analysed the expression of the CX3CL1 and CX3CR1 mRNA and protein in the duodenal biopsies of CD patients with active and treated disease, and in non-CD control individuals, by qPCR and immunohistochemistry. Results: After the gluten challenge, increased levels of CX3CL1, I-TAC and MIG proteins were observed in the peripheral blood of CD patients, with no changes in CX3CL1 mRNA, or CX3CR1 mRNA and protein. Regarding duodenal tissue, CX3CL1 was absent or barely present in the superficial and basal epithelium of CD patients, contrasting with the moderate to high levels present in controls. Conclusions: CX3CL1 seems to be involved in the appearance and progression of CD, and it appears to be a potential diagnostic biomarker. Its use as an alternative therapeutic target in CD deserves further research.

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Flow cytometric discrimination of seven lineage markers by using two fluorochromes

Cited by 4 publications

References 19 publications

Onchocerca volvulus bivalent subunit vaccine induces protective immunity in genetically diverse collaborative cross recombinant inbred intercross mice

Onchocerca volvulus bivalent subunit vaccine induces protective immunity in genetically diverse collaborative cross recombinant inbred intercross mice

Role of crotoxin in coagulation: novel insights into anticoagulant mechanisms and impairment of inflammation-induced coagulation

CX3CL1–CX3CR1 Axis: A New Player in Coeliac Disease Pathogenesis

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