rabbit anti d5r  (Alomone Labs)

Journal: Immunology

Article Title: Instantaneous depolarization of T cells via dopamine receptors, and inhibition of activated T cells of Psoriasis patients and inflamed human skin, by D1‐like receptor agonist: Fenoldopam

doi: 10.1111/imm.13109

Figure Lengend Snippet: Dopamine receptor agonists (100 n m ) by themselves shift the membrane potential ( V m ) of normal human T cells, within 15 seconds only.

Article Snippet: In single‐staining experiments, normal human CD3 + T cells (isolated from fresh human blood samples of healthy blood donors) were stained with rabbit‐anti‐human D1R, D2R, D3R, D4R and D5R polyclonal antibodies (Calbiochem, http://www.merckmillipore.com/ ) (1 : 50), or with rabbit isotype control (1 : 50), and then with fluorescein isothiocyanate (FITC)‐conjugated goat anti‐rabbit antibody (Jackson Laboratories, https://www.jacksonimmuno.com ) (1 : 100).

Techniques: Membrane, Control

Journal: Immunology

Article Title: Instantaneous depolarization of T cells via dopamine receptors, and inhibition of activated T cells of Psoriasis patients and inflamed human skin, by D1‐like receptor agonist: Fenoldopam

doi: 10.1111/imm.13109

Figure Lengend Snippet: Selective D1/5R agonists induce depolarization of activated CD3+ normal human T cells within 15 seconds. (a–g) The fluorescence profiles of the voltage‐sensitive fluorescence dye DiBac3 inside activated normal human T cells, both before (gray) and then again 15 seconds after (colored) addition of either: Control buffer (PBS) only (a), D1R/D15R (D1‐like) agonists: SKF 38393 (b) and Fenoldopam (c); D2R agonist: Sumanirole (d); D3R agonists: 7‐OH DPAT (e) and PD 128907 (f); or D4R agonist: PD 168077 (g). All the DR agonists were used in a final concentration of 10−7 M (100 nM), prepared freshly before each experiment, from either their powders or very concentrated stocks stored at −80°. The corresponding intensity of the voltage‐sensitive fluorescence dye DiBac3 was determined by the geometric mean (GM) of the fluorescence (FL1), in each tube before and after addition of each agonist, and all the GMs and fold change/shift are shown in Table 1. As for the depolarization of resting T cells shown in Fig 2, the results of the activated T cells shown in the figure are of one representative experiment, of two performed altogether, on T cells of different healthy human paticipants. In both experiments the same pattern of results was observed, but the actual extent of the depolarization varied. This was not surprising in view of the differences between individuals with regard to the level of DRs on the cell surface of their T cells found in7 and also in the present study (data not shown).

Article Snippet: In single‐staining experiments, normal human CD3 + T cells (isolated from fresh human blood samples of healthy blood donors) were stained with rabbit‐anti‐human D1R, D2R, D3R, D4R and D5R polyclonal antibodies (Calbiochem, http://www.merckmillipore.com/ ) (1 : 50), or with rabbit isotype control (1 : 50), and then with fluorescein isothiocyanate (FITC)‐conjugated goat anti‐rabbit antibody (Jackson Laboratories, https://www.jacksonimmuno.com ) (1 : 100).

Techniques: Fluorescence, Control, Concentration Assay

Journal: Immunology

Article Title: Instantaneous depolarization of T cells via dopamine receptors, and inhibition of activated T cells of Psoriasis patients and inflamed human skin, by D1‐like receptor agonist: Fenoldopam

doi: 10.1111/imm.13109

Figure Lengend Snippet: Selective D1/5R agonists induce depolarization of resting CD3+ normal human T cells within 15 seconds. (a–g) The fluorescence profiles of the voltage‐sensitive fluorescence dye DiBac3 inside resting normal human T cells, both before (gray) and then again 15 seconds after (colored) addition of either: Control buffer (PBS) only (a), D1R/D15R (D1‐like) agonists: SKF 38393 (b) and Fenoldopam (c); D2R agonist: Sumanirole (d); D3R agonists: 7‐OH DPAT (e) and PD 128907 (f); D4R agonist: PD 168077 (g). All the DR agonists were used in a final concentration of 10−7 M (100 nM), prepared freshly before each experiment, from either their powders or very concentrated stocks stored in at –80°. The corresponding intensity of the voltage‐sensitive fluorescence dye DiBac3 was determined by the geometric mean (GM) of the fluorescence (FL1), in each tube before and after addition of each agonist, and all the GMs and fold change/shift are shown in Table 1. The results shown in the figure are of one representative experiment, of two performed altogether, on T cells of different healthy human participants. In both experiments the same pattern of results was observed, but the actual extent of the depolarization varied. This was not surprising in view of the differences between individuals with regard to the level of DRs on the cell surface of their T cells (found in 7 and also in the current study (data not shown).

Article Snippet: In single‐staining experiments, normal human CD3 + T cells (isolated from fresh human blood samples of healthy blood donors) were stained with rabbit‐anti‐human D1R, D2R, D3R, D4R and D5R polyclonal antibodies (Calbiochem, http://www.merckmillipore.com/ ) (1 : 50), or with rabbit isotype control (1 : 50), and then with fluorescein isothiocyanate (FITC)‐conjugated goat anti‐rabbit antibody (Jackson Laboratories, https://www.jacksonimmuno.com ) (1 : 100).

Techniques: Fluorescence, Control, Concentration Assay

Journal: Immunology

Article Title: Instantaneous depolarization of T cells via dopamine receptors, and inhibition of activated T cells of Psoriasis patients and inflamed human skin, by D1‐like receptor agonist: Fenoldopam

doi: 10.1111/imm.13109

Figure Lengend Snippet: Selective D1/5R agonists induce depolarization of activated CD3+ normal human T cells within 15 seconds. (a–g) The fluorescence profiles of the voltage‐sensitive fluorescence dye DiBac3 inside activated normal human T cells, both before (gray) and then again 15 seconds after (colored) addition of either: Control buffer (PBS) only (a), D1R/D15R (D1‐like) agonists: SKF 38393 (b) and Fenoldopam (c); D2R agonist: Sumanirole (d); D3R agonists: 7‐OH DPAT (e) and PD 128907 (f); or D4R agonist: PD 168077 (g). All the DR agonists were used in a final concentration of 10−7 M (100 nM), prepared freshly before each experiment, from either their powders or very concentrated stocks stored at −80°. The corresponding intensity of the voltage‐sensitive fluorescence dye DiBac3 was determined by the geometric mean (GM) of the fluorescence (FL1), in each tube before and after addition of each agonist, and all the GMs and fold change/shift are shown in Table 1. As for the depolarization of resting T cells shown in Fig 2, the results of the activated T cells shown in the figure are of one representative experiment, of two performed altogether, on T cells of different healthy human paticipants. In both experiments the same pattern of results was observed, but the actual extent of the depolarization varied. This was not surprising in view of the differences between individuals with regard to the level of DRs on the cell surface of their T cells found in7 and also in the present study (data not shown).

Article Snippet: In single‐staining experiments, normal human CD3 + T cells (isolated from fresh human blood samples of healthy blood donors) were stained with rabbit‐anti‐human D1R, D2R, D3R, D4R and D5R polyclonal antibodies (Calbiochem, http://www.merckmillipore.com/ ) (1 : 50), or with rabbit isotype control (1 : 50), and then with fluorescein isothiocyanate (FITC)‐conjugated goat anti‐rabbit antibody (Jackson Laboratories, https://www.jacksonimmuno.com ) (1 : 100).

Techniques: Fluorescence, Concentration Assay

Journal: Immunology

Article Title: Instantaneous depolarization of T cells via dopamine receptors, and inhibition of activated T cells of Psoriasis patients and inflamed human skin, by D1‐like receptor agonist: Fenoldopam

doi: 10.1111/imm.13109

Figure Lengend Snippet: Selective D1/5R agonists induce depolarization of resting CD3+ normal human T cells within 15 seconds. (a–g) The fluorescence profiles of the voltage‐sensitive fluorescence dye DiBac3 inside resting normal human T cells, both before (gray) and then again 15 seconds after (colored) addition of either: Control buffer (PBS) only (a), D1R/D15R (D1‐like) agonists: SKF 38393 (b) and Fenoldopam (c); D2R agonist: Sumanirole (d); D3R agonists: 7‐OH DPAT (e) and PD 128907 (f); D4R agonist: PD 168077 (g). All the DR agonists were used in a final concentration of 10−7 M (100 nM), prepared freshly before each experiment, from either their powders or very concentrated stocks stored in at –80°. The corresponding intensity of the voltage‐sensitive fluorescence dye DiBac3 was determined by the geometric mean (GM) of the fluorescence (FL1), in each tube before and after addition of each agonist, and all the GMs and fold change/shift are shown in Table 1. The results shown in the figure are of one representative experiment, of two performed altogether, on T cells of different healthy human participants. In both experiments the same pattern of results was observed, but the actual extent of the depolarization varied. This was not surprising in view of the differences between individuals with regard to the level of DRs on the cell surface of their T cells (found in 7 and also in the current study (data not shown).

Article Snippet: In single‐staining experiments, normal human CD3 + T cells (isolated from fresh human blood samples of healthy blood donors) were stained with rabbit‐anti‐human D1R, D2R, D3R, D4R and D5R polyclonal antibodies (Calbiochem, http://www.merckmillipore.com/ ) (1 : 50), or with rabbit isotype control (1 : 50), and then with fluorescein isothiocyanate (FITC)‐conjugated goat anti‐rabbit antibody (Jackson Laboratories, https://www.jacksonimmuno.com ) (1 : 100).

Techniques: Fluorescence, Concentration Assay

Journal: Immunology

Article Title: Instantaneous depolarization of T cells via dopamine receptors, and inhibition of activated T cells of Psoriasis patients and inflamed human skin, by D1‐like receptor agonist: Fenoldopam

doi: 10.1111/imm.13109

Figure Lengend Snippet: Normal human CD3+ T cells express all types of dopamine receptors (DRS) – D1R‐D5R on their cell surface, and the level of D1R, D5R and D4R are higher in activated T cells than in resting T cells. (a–e; Upper Figs) Expression of D1R–D5R on resting normal human CD3+ T cells. (f–j; Lower Figs) Expression of D1R–D5R on CD3/CD28‐activated normal human CD3+ T cells. The colored graphs show the levels of immunofluorescence staining, first with rabbit‐anti‐human antibodies selective to each DR type, and then with a secondary‐FITC‐conjugated goat anti‐rabbit antibody. The control gray fluorescence profiles in each graph represent the control non‐specific staining, first with rabbit isotype control, and then with the same secondary antibody. The results are of one representative experiment, of two performed on T cells of different healthy human individuals. In both experiments the same pattern of results was observed, but the actual numbers/expression levels were different. This observation is in line with the findings of Kustrimovic et al.,7 that human T cells express all DR types, but that there is a remarkable variability between different people with regard to the exact level of each DR type expressed on the cell surface of their T cells.

Article Snippet: In single‐staining experiments, normal human CD3 + T cells (isolated from fresh human blood samples of healthy blood donors) were stained with rabbit‐anti‐human D1R, D2R, D3R, D4R and D5R polyclonal antibodies (Calbiochem, http://www.merckmillipore.com/ ) (1 : 50), or with rabbit isotype control (1 : 50), and then with fluorescein isothiocyanate (FITC)‐conjugated goat anti‐rabbit antibody (Jackson Laboratories, https://www.jacksonimmuno.com ) (1 : 100).

Techniques: Expressing, Immunofluorescence, Staining, Fluorescence