human cd80 pe conjugated antibody  (R&D Systems)

Journal: ACS Biomaterials Science & Engineering

Article Title: Methods and Technical Issues for Optimizing the Production of Hydrogels Containing Decellularized Wharton’s Jelly

doi: 10.1021/acsbiomaterials.5c02006

Figure Lengend Snippet: Human M1-polarized macrophages were exposed to A075G3 and A075W3 for 48 h. Positivity for CD80, a pro-inflammatory surface marker, was assessed by flow cytometry and expressed as mean fluorescence intensity (MFI) ± SD ( n = 4 for CTR, A075G3 and A075W3). * p < 0.05 (A075W3 vs CTR).

Article Snippet: PE conjugated antihuman CD80 Antibody (clone REA661) from Miltenyi Biotec (Germany).

Techniques: Marker, Flow Cytometry, Fluorescence

Journal: Frontiers in Immunology

Article Title: Cladribine Treatment for MS Preserves the Differentiative Capacity of Subsequently Generated Monocytes, Whereas Its Administration In Vitro Acutely Influences Monocyte Differentiation but Not Microglial Activation

doi: 10.3389/fimmu.2022.678817

Figure Lengend Snippet: Effects of oral cladribine treatment on circulating PBMCs from MS patients and monocyte ex vivo differentiation. Monocytes isolated from MS patients’ PBMCs before (white) and 19-21 days after (blue) the beginning of cladribine treatment were differentiated to MDMs for 7 days with M-CSF. (A) The density of peripheral blood mononuclear cells (PBMC) in blood collected from MS patients before (BL; baseline) and 19-21 days after (FU; follow up) the beginning of cladribine treatment. (B) The density of MACS-isolated monocytes from freshly isolated PBMCs of MS patients before (BL; baseline) and 19-21 days after (FU; follow up) the beginning of cladribine treatment. (C) Monocyte subsets from cladribine-treated MS patients before (white) and 19-21 days after (blue) the beginning of cladribine treatment were determined by CD14 and CD16 surface expression, as analyzed by flow cytometry. (D) MDMs of cladribine-treated MS patients were differentiated for 7 days with M-CSF and the expression of pro-inflammatory (CD80) and anti-inflammatory (CD163 and MERTK) activation markers were analyzed by flow cytometry. (E–I) Gene expression of activation markers in monocyte-derived macrophages (MDM) from cladribine-treated MS patients. RT-qPCR gene expression analysis of pro-inflammatory (E–G) and anti-inflammatory (H, I) activation markers. Positive cells were determined according to (C) isotype controls and (D) FMO controls. (A–D) Data depicted as mean (with SD) and lines indicate before-after cladribine treatment, (A, C) n=9, (B–I) n=10. The yellow crossed diamond shows the statistically significant outlier (ESD, extreme studentized deviate method), which was excluded from the analysis. (A–D) Statistical significance was calculated with paired t-test. (E–I) Data depicted as median (with quartiles, dot indicates mean), lines indicate before-after cladribine treatment, n=9 (patient #9 was not included due to technical problems). (E) n=8, the outlier value #7 follow up (ESD method) was not included in the analysis. (E–I) p-values between baseline and follow up were determined by paired t-test and the adjusted p-value (p adj ) was calculated using the posthoc Holm-Sidak correction for multiple testing.

Article Snippet: Blocked cells were stained for 15 min at RT with anti-human CD80 PE-conjugated antibody (clone REA661; Miltenyi, #130-110-270), anti-human CD163 VioBlue-conjugated antibody (clone REA812; Miltenyi #130-112-134) and anti-human Mer APC-conjugated antibody (R&D SYSTEMS, #FAB8912A) at a final dilution of 1:100, 1:50 and 1:11, respectively.

Techniques: Ex Vivo, Isolation, Expressing, Flow Cytometry, Activation Assay, Gene Expression, Derivative Assay, Quantitative RT-PCR

Journal: Frontiers in Immunology

Article Title: Cladribine Treatment for MS Preserves the Differentiative Capacity of Subsequently Generated Monocytes, Whereas Its Administration In Vitro Acutely Influences Monocyte Differentiation but Not Microglial Activation

doi: 10.3389/fimmu.2022.678817

Figure Lengend Snippet: Expression of activation markers in monocyte-derived macrophages (MDMs) treated with cladribine in vitro . Monocytes isolated from healthy donors were differentiated to MDMs with M-CSF and/or GM-CSF in the presence of cladribine. (A–C) Percentage of MDMs expressing pro-inflammatory (CD80) and anti-inflammatory (CD163 and MERTK) activation markers at day 6 of cladribine treatment in vitro as analyzed by flow cytometry. Positive cells were determined according to their fluorescent minus one (FMO) control. (D) Levels of CD80 surface protein expression on MDMs treated with cladribine during differentiation in vitro , as analyzed by flow cytometry. (E–I) RT-qPCR gene expression analysis of (E–G) pro-inflammatory and (H, I) anti-inflammatory activation markers at day 6 post cladribine treatment in vitro . (A–C) Positive cells were determined according to FMO controls. (A–D) Data depicted as mean (with SD) and symbols indicate matched experiments from the same healthy donor’s sample, n=2. p-values between cladribine 0.05 μM and vehicle (DMSO) were determined by ordinary two-way ANOVA with Sidak’s posthoc test. (E–I) Data depicted as mean (with max-min) and symbols indicate matched experiments from the same healthy donor’s sample, n=3. p-values between cladribine 0.05 μM and vehicle (DMSO) were determined by ordinary two-way ANOVA with Sidak’s posthoc test.

Article Snippet: Blocked cells were stained for 15 min at RT with anti-human CD80 PE-conjugated antibody (clone REA661; Miltenyi, #130-110-270), anti-human CD163 VioBlue-conjugated antibody (clone REA812; Miltenyi #130-112-134) and anti-human Mer APC-conjugated antibody (R&D SYSTEMS, #FAB8912A) at a final dilution of 1:100, 1:50 and 1:11, respectively.

Techniques: Expressing, Activation Assay, Derivative Assay, In Vitro, Isolation, Flow Cytometry, Control, Quantitative RT-PCR, Gene Expression