Integrated measurements by flow cytometry of the cytokines IL-2, IFN-γ, IL-12, TNF-α and functional evaluation of their receptors in human blood

Adriana Garibay-Escobar, Iris Estrada-García, Sergio Estrada-Parra, Leopoldo Santos-Argumedo

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Abstract

Immunodeficiencies might be caused not only by the lack of cytokine production, but also by defective expression and/or function of the cytokine receptors. We have measured by flow cytometry, within 2 days, not only the production of IL-2, IFN-γ, IL-12 and TNF-α, but also the functional expression of the receptors for these cytokines in blood samples obtained from 15 healthy donors and 13 patients suffering from tuberculosis. Cytoplasmic and surface staining with monoclonal antibodies (mAbs) was used to assess the production of cytokines and their receptors, respectively, after polyclonal stimulation. To evaluate receptor activity, peripheral blood mononuclear cells (PBMC) were first incubated with the corresponding recombinant human (rh) cytokine. CD69 was detected on lymphocytes after incubation with rhIL-2; IFN-γ was detected in lymphocytes after co-stimulation with rhIL-12 plus PHA; iNOS induction and upregulation of major histocompatibility complex (MHC) II and MHC I was detected on monocytes after recombinant human interferon-γ (rhIFN-γ) stimulation; finally, COX-2 expression and MHC II upregulation were detected on monocytes after rhTNF-α stimulation. The assay that was developed can be used clinically to assess the activities of components of the cytokine signaling pathways of patients with immunodeficiencies or those with chronic intracellular infections such as tuberculosis. © 2003 Elsevier B.V. All rights reserved.
Original languageAmerican English
Pages (from-to)73-88
Number of pages64
JournalJournal of Immunological Methods
DOIs
StatePublished - 1 Sep 2003

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Cytokine Receptors
cytometry
Flow cytometry
Interleukin-12
Major Histocompatibility Complex
stimulation
blood
Interleukin-2
Flow Cytometry
Blood
Lymphocytes
monocytes
Cytokines
tuberculosis
evaluation
Monocytes
lymphocytes
Tuberculosis
Up-Regulation
interferon

Cite this

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title = "Integrated measurements by flow cytometry of the cytokines IL-2, IFN-γ, IL-12, TNF-α and functional evaluation of their receptors in human blood",
abstract = "Immunodeficiencies might be caused not only by the lack of cytokine production, but also by defective expression and/or function of the cytokine receptors. We have measured by flow cytometry, within 2 days, not only the production of IL-2, IFN-γ, IL-12 and TNF-α, but also the functional expression of the receptors for these cytokines in blood samples obtained from 15 healthy donors and 13 patients suffering from tuberculosis. Cytoplasmic and surface staining with monoclonal antibodies (mAbs) was used to assess the production of cytokines and their receptors, respectively, after polyclonal stimulation. To evaluate receptor activity, peripheral blood mononuclear cells (PBMC) were first incubated with the corresponding recombinant human (rh) cytokine. CD69 was detected on lymphocytes after incubation with rhIL-2; IFN-γ was detected in lymphocytes after co-stimulation with rhIL-12 plus PHA; iNOS induction and upregulation of major histocompatibility complex (MHC) II and MHC I was detected on monocytes after recombinant human interferon-γ (rhIFN-γ) stimulation; finally, COX-2 expression and MHC II upregulation were detected on monocytes after rhTNF-α stimulation. The assay that was developed can be used clinically to assess the activities of components of the cytokine signaling pathways of patients with immunodeficiencies or those with chronic intracellular infections such as tuberculosis. {\circledC} 2003 Elsevier B.V. All rights reserved.",
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T1 - Integrated measurements by flow cytometry of the cytokines IL-2, IFN-γ, IL-12, TNF-α and functional evaluation of their receptors in human blood

AU - Garibay-Escobar, Adriana

AU - Estrada-García, Iris

AU - Estrada-Parra, Sergio

AU - Santos-Argumedo, Leopoldo

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N2 - Immunodeficiencies might be caused not only by the lack of cytokine production, but also by defective expression and/or function of the cytokine receptors. We have measured by flow cytometry, within 2 days, not only the production of IL-2, IFN-γ, IL-12 and TNF-α, but also the functional expression of the receptors for these cytokines in blood samples obtained from 15 healthy donors and 13 patients suffering from tuberculosis. Cytoplasmic and surface staining with monoclonal antibodies (mAbs) was used to assess the production of cytokines and their receptors, respectively, after polyclonal stimulation. To evaluate receptor activity, peripheral blood mononuclear cells (PBMC) were first incubated with the corresponding recombinant human (rh) cytokine. CD69 was detected on lymphocytes after incubation with rhIL-2; IFN-γ was detected in lymphocytes after co-stimulation with rhIL-12 plus PHA; iNOS induction and upregulation of major histocompatibility complex (MHC) II and MHC I was detected on monocytes after recombinant human interferon-γ (rhIFN-γ) stimulation; finally, COX-2 expression and MHC II upregulation were detected on monocytes after rhTNF-α stimulation. The assay that was developed can be used clinically to assess the activities of components of the cytokine signaling pathways of patients with immunodeficiencies or those with chronic intracellular infections such as tuberculosis. © 2003 Elsevier B.V. All rights reserved.

AB - Immunodeficiencies might be caused not only by the lack of cytokine production, but also by defective expression and/or function of the cytokine receptors. We have measured by flow cytometry, within 2 days, not only the production of IL-2, IFN-γ, IL-12 and TNF-α, but also the functional expression of the receptors for these cytokines in blood samples obtained from 15 healthy donors and 13 patients suffering from tuberculosis. Cytoplasmic and surface staining with monoclonal antibodies (mAbs) was used to assess the production of cytokines and their receptors, respectively, after polyclonal stimulation. To evaluate receptor activity, peripheral blood mononuclear cells (PBMC) were first incubated with the corresponding recombinant human (rh) cytokine. CD69 was detected on lymphocytes after incubation with rhIL-2; IFN-γ was detected in lymphocytes after co-stimulation with rhIL-12 plus PHA; iNOS induction and upregulation of major histocompatibility complex (MHC) II and MHC I was detected on monocytes after recombinant human interferon-γ (rhIFN-γ) stimulation; finally, COX-2 expression and MHC II upregulation were detected on monocytes after rhTNF-α stimulation. The assay that was developed can be used clinically to assess the activities of components of the cytokine signaling pathways of patients with immunodeficiencies or those with chronic intracellular infections such as tuberculosis. © 2003 Elsevier B.V. All rights reserved.

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