Detection by immunofluorescence of surface molecules present in low copy numbers high sensitivity staining and calibration of flow cytometer

Zola, Heddy; Neoh, S. H.; Mantzioris, Basil X.; Webster, Joseph B.; Loughnan, Michael

doi:10.1016/0022-1759(90)90278-4

Cited by 65 publications

(32 citation statements)

References 17 publications

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“…Several possibilities could account for an accumulation of CD4 memory T cells in the absence of detectable T cell activation, at a site of invasion by a pathogen: (i) only those T cells specific for mycobacterial antigens are activated and, as PPDresponder frequencies have been estimated for tuberculous effusions as being in the region of 1/1000 to 1/7000 [2], this level of T cell activation would not be detectable [18,19]; (ii) the effusion may not be the site of pathology, but rather the adjacent tissue. T cells in the effusion could be ineffective spill-over cells not actively involved in combating the infection; (iii) regulatory mechanisms may be active in the effusion which limit T cell responses, perhaps to minimize tissue damage while allowing effective pathogen control.…”

Section: Discussionmentioning

confidence: 99%

Memory lymphocytes from tuberculous effusions: purified protein derivative (PPD) stimulates accelerated activation marker expression and cell cycle progression

Lukey

Latouf

Ress

1996

Clinical and Experimental Immunology

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SUMMARYAccelerated PPD-specific proliferation and generation of CD4 cytotoxic effectors by mononuclear leucocytes (MNL) from tuberculous effusions (EMNL) has been previously reported by our laboratory. In order to explore the contribution of the state of activation of MNL to accelerated reactivity, EMNL and peripheral blood (PB)MNL from seven patients with tuberculosis were assessed both ex vivo and after PPD stimulation. Flow cytometry revealed no difference in the activation state (IL-2 receptor and HLA-DR expression) or cell cycle progression ex vivo. However, CD4 CD29 memory T cells were accumulated in EMNL compared with PBMNL. In vitro stimulation of EMNL with PPD resulted in accelerated expression of activation markers and progression through the cell cycle (peak after 4 days), whilst PBMNL exhibited normal activation kinetics (peak after 7 days). Accelerated reactivity could not be accounted for by quantitative differences in effusion CD4 CD29 memory T cells compared with blood, but may be due to a qualitative difference in effusion memory T cells, which are shown to be in a post-activation state of differentiation. T cells entering S and G 2 /M phases of the cell cycle were largely of the activated memory phenotype. Activation marker expression occurred in association with upregulation of CD4 antigen expression on the surface of EMNL. Thus accelerated expression of activation markers and cell cycle progression by CD4 CD29 memory T cells may in part account for accelerated PPD reactivity in tuberculous effusions.

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Section: Discussionmentioning

confidence: 99%

Memory lymphocytes from tuberculous effusions: purified protein derivative (PPD) stimulates accelerated activation marker expression and cell cycle progression

Lukey

Latouf

Ress

1996

Clinical and Experimental Immunology

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“…Flow cytometry experiments using an unconjugated antibody were performed using a three-step "high sensitivity" staining protocol [21]. Briefly, PBMC were incubated with In experiments where antibodies directed against intracellular antigens were used, the surface staining as described above was performed first, and the cells were then permeabilised and stained according to the manufacturer's instructions.…”

Section: 4flow Cytometrymentioning

confidence: 99%

PI16 is expressed by a subset of human memory Treg with enhanced migration to CCL17 and CCL20

Nicholson

Mavrangelos

Bird

et al. 2012

Cellular Immunology

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“…We observed that, in most experiments, the amount of membrane mAb molecules on our effectors was in the nanomolar range, which is far below the dose usually recommended to obtain redirected cytotoxicity. Furthermore, with a more sensitive assay that has been reported to detect as few as 400 molecules/ cell [65], no additional BMA030 was detected in any of the experiments (results not illustrated).…”

Section: Comparison Of Oc/tr Bispecific Mab Retargeting Of T Cells Stmentioning

confidence: 86%

Further characterization of cytotoxic T cells generated by short-term culture of human peripheral blood lymphocytes with interleukin-2 and anti-CD3 mAb

Jacobs

Greimers

Mazzoni

et al. 1996

Cancer Immunology, Immunotherapy

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Detection by immunofluorescence of surface molecules present in low copy numbers high sensitivity staining and calibration of flow cytometer

Cited by 65 publications

References 17 publications

Memory lymphocytes from tuberculous effusions: purified protein derivative (PPD) stimulates accelerated activation marker expression and cell cycle progression

Memory lymphocytes from tuberculous effusions: purified protein derivative (PPD) stimulates accelerated activation marker expression and cell cycle progression

PI16 is expressed by a subset of human memory Treg with enhanced migration to CCL17 and CCL20

Further characterization of cytotoxic T cells generated by short-term culture of human peripheral blood lymphocytes with interleukin-2 and anti-CD3 mAb

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