Journal: Frontiers in Immunology

Article Title: Design and validation of novel flow cytometry panels to analyze a comprehensive range of peripheral immune cells in mice

doi: 10.3389/fimmu.2024.1432816

Figure Lengend Snippet: Antibodies used in the myeloid and lymphoid panels.

Article Snippet: CD172a , VioBlue , Miltenyi , 0.5μL , 130-123-151 , REA1201.

Techniques:

Journal: Frontiers in Immunology

Article Title: Design and validation of novel flow cytometry panels to analyze a comprehensive range of peripheral immune cells in mice

doi: 10.3389/fimmu.2024.1432816

Figure Lengend Snippet: Intracellular panel at 72 hours post-injection (p.i.). This figure shows relative counts from the Intracellular panel in animals treated with LPS and in control animals at 72 hours post-injection. Only populations with significant differences between groups are presented. Significance levels are indicated as follows: *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001. (A) CD45 Leukocytes; (B) Monocyte type myeloid derived suppressor cell; (C) Lymphocytes; (D) Tcells; (E) CD4 Tcells; (F) CD4 Effector memory; (G) CD4 CD44 positive; (H) T Helper 9 Cells; (I) CD8 Tcells; (J) CD8 Central memory; (K) CD8 Effector activated; (L) lymphocytes B; (M) Natural Killers; (N) CD8 Effector memory; (O) plasma cells; (P) Natural killer T cell; (Q) CD172a myeloids cells.

Article Snippet: CD172a , VioBlue , Miltenyi , 0.5μL , 130-123-151 , REA1201.

Techniques: Injection, Control, Derivative Assay, Clinical Proteomics

Journal: Frontiers in Immunology

Article Title: Design and validation of novel flow cytometry panels to analyze a comprehensive range of peripheral immune cells in mice

doi: 10.3389/fimmu.2024.1432816

Figure Lengend Snippet: Proportions of variance (ƞ2) associated with intra- and inter-individual factors.

Article Snippet: CD172a , VioBlue , Miltenyi , 0.5μL , 130-123-151 , REA1201.

Techniques: Clinical Proteomics

Journal: Frontiers in Immunology

Article Title: Design and validation of novel flow cytometry panels to analyze a comprehensive range of peripheral immune cells in mice

doi: 10.3389/fimmu.2024.1432816

Figure Lengend Snippet: Effect size corresponding to a Cohen’s d of 0.5 for the overall variables studied in the myeloid and lymphoid panels.

Article Snippet: CD172a , VioBlue , Miltenyi , 0.5μL , 130-123-151 , REA1201.

Techniques: Clinical Proteomics

Journal: Scientific Reports

Article Title: Role of Signal Regulatory Protein α in Arsenic Trioxide-induced Promyelocytic Leukemia Cell Apoptosis

doi: 10.1038/srep23710

Figure Lengend Snippet: ( a ) Western blotting of p-Akt, Akt, p-GSK-3β, GSK-3β, β-catenin, Foxo3a and GAPDH in HL-60 and Huh7 cells infected with LV-SIRPα or LV-CTL on the third day post infection: representative Western blotting (left panels) and quantitative analysis (right panels). ( b ) Western blotting of β-catenin, Foxo3a, cleaved caspase-3 and GAPDH in LV-SIRPα-infected HL-60 cells co-incubated with 0, 5 and 10 μM LiCl or 0, 5, or 10 mM SB-216763. 48 hours post-infection of lentivirus, cells were treated with LiCl, or SB-216763 for 4 h and then lysed for Western blotting analysis: representative Western blot (left panels) and quantitative analysis (right panels). Values were shown as the mean ± SEM (n = 3). *P < 0.05. **P < 0.01.

Article Snippet: Antibodies against GAPDH, SIRPα, and secondary antibodies against mouse or rabbit IgGs were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Western Blot, Infection, Incubation

Journal: Scientific Reports

Article Title: Role of Signal Regulatory Protein α in Arsenic Trioxide-induced Promyelocytic Leukemia Cell Apoptosis

doi: 10.1038/srep23710

Figure Lengend Snippet: ( a,b ) Western blotting of p-Akt, Akt, p-GSK-3β, GSK-3β, β-catenin, Foxo3a and GAPDH in HL-60 or NB4 cells treated with ATO for indicated time: representative Western blotting ( a ) and quantitative analysis of protein level ( b ), the protein level of GAPDH was used as an internal control, the p-Akt (Ser473) and p-GSK-3β (Ser9) were normalized to the total Akt and GSK-3β, respectively. ( c,d ) Western blotting of p-Akt, Akt, p-GSK-3β, GSK-3β, β-catenin, Foxo3a and GAPDH in SIRPα shRNA lentivirus infected HL-60 or NB4 cells after the treatment of ATO for 48 h: representative Western blot ( c ) and quantitative analysis ( d ). Values were shown as the mean ± SEM (n = 3). *P < 0.05. **P < 0.01.

Article Snippet: Antibodies against GAPDH, SIRPα, and secondary antibodies against mouse or rabbit IgGs were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Western Blot, Control, shRNA, Infection

Journal: Scientific Reports

Article Title: Role of Signal Regulatory Protein α in Arsenic Trioxide-induced Promyelocytic Leukemia Cell Apoptosis

doi: 10.1038/srep23710

Figure Lengend Snippet: ( a ) The level of SIRPα-regulating miRNAs, miR-17, miR-20a, miR-106a in HL-60 or NB4 cells treated with ATO at indicated time. The level of all miRNAs was normalized to that of U6. ( b ) The relative mRNA level of SIRPα in HL-60 or NB4 cells treated with ATO for indicated time. Total RNA was extracted from the cells and analyzed with RT-qPCR. The mRNA level of GAPDH was used as an internal control. ( c ) Western blotting of SIRPα protein level in the HL-60 or NB4 cells treated with ATO for indicated time. Before ATO treatment, the cells were transfected with pre-miR-17. The mock-transfected cells (PBS) or cells transfected with scrambles oligonucleotide were used as a control: representative Western blot (upper panel) and quantitative analysis (lower panel). ( d ) Flow cytometry analysis of apoptosis of pre-miR-17-transfected HL-60 or NB4 cells after the treatment of ATO for 48 h: representative flow cytometer data (left panel) and quantitative analysis of apoptosis (right panel). The percentage of annexin V-positive cells was calculated. Values were shown as the mean ± SEM (n = 3). *P < 0.05. **P < 0.01.

Article Snippet: Antibodies against GAPDH, SIRPα, and secondary antibodies against mouse or rabbit IgGs were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Quantitative RT-PCR, Control, Western Blot, Transfection, Flow Cytometry

Journal: Journal of Cosmetic Dermatology

Article Title: Elevated CD47 Expression Impairs Elimination of Photoaged Fibroblasts by Macrophages and Serves as a Potential Biomarker for Photoaging

doi: 10.1111/jocd.70098

Figure Lengend Snippet: The CD47‐SIRPα axis impairs macrophage‐mediated phagocytosis and blockage of CD47‐SIRPα axis could improve the clearance of photoaged fibroblasts by macrophages. (A) Western blotting for SIRPα expression in macrophages co‐cultured with photoaged fibroblasts. (B) Representative pictures of SIRPα immunofluorescence staining. Red fluorescence for SIRPα, blue fluorescence for DAPI. Scale bar, 25 μm. (C) Macrophages were first co‐cultured with control or photoaged fibroblasts for 16 h and then switched to co‐culture with CFSE‐labeled (green) apoptotic Jurkat cells for 2 h. Representative images and quantified percentages of CFSE‐labeled (green fluorescence) apoptotic Jurkat cells phagocytosed by macrophages in each group. Scale bar, 50 μm. (D) Western blot analysis and quantification analysis of CD47 protein expression in photoaged fibroblasts transfected with CD47 siRNA or control siRNA. (E) Western blot analysis and quantification analysis of SIRPα protein expression in macrophages transfected with SIRPα siRNA or control siRNA. (F) Representative images of the photoaged fibroblasts‐macrophages co‐culture system with indicated treatments and related quantitative analysis of photoaged fibroblasts counts. The green color represents the residual CFSE‐labeled photoaged fibroblasts un‐eliminated by macrophages. Scale bar, 50 μm. Data are the means ± SD from three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001. FBs, fibroblasts. Mφ, macrophage.

Article Snippet: To determine the role of CD47‐SIRPα axis in this process, CD47 siRNA (sc‐35 006, Santa Cruz, CA, USA) and SIRPα siRNA (sc‐44 106, Santa Cruz) were applied to block their expression using Lipo3000 (Invitrogen, CA, USA), respectively.

Techniques: Western Blot, Expressing, Cell Culture, Immunofluorescence, Staining, Fluorescence, Control, Co-Culture Assay, Labeling, Transfection

Journal: Journal of Cosmetic Dermatology

Article Title: Elevated CD47 Expression Impairs Elimination of Photoaged Fibroblasts by Macrophages and Serves as a Potential Biomarker for Photoaging

doi: 10.1111/jocd.70098

Figure Lengend Snippet: Increased number of CD47 + senescent fibroblasts and SIRPα + macrophages in the sun‐exposed aged skin. (A) Triple immunofluorescence staining for CD47 (deep red), Vimentin (green) and p21 (red) in skin tissues of sun‐protected and sun‐exposed groups. Blue: DAPI nuclear counterstaining. Bar graphs represent the number of CD47, p16, and Vimentin triple‐positive cells. Scale bar, 50 μm. (B) Double immunofluorescence staining for CD68 (green) and SIRPα (red) in skin tissues of each group. Blue: DAPI nuclear counterstaining. Bar graphs represent the number of CD68/SIRPα double‐positive cells. Data are the means ± SD from three independent experiments. *** p < 0.001. FBs, fibroblasts. Mφ, macrophage.

Article Snippet: To determine the role of CD47‐SIRPα axis in this process, CD47 siRNA (sc‐35 006, Santa Cruz, CA, USA) and SIRPα siRNA (sc‐44 106, Santa Cruz) were applied to block their expression using Lipo3000 (Invitrogen, CA, USA), respectively.

Techniques: Immunofluorescence, Staining, Double Immunofluorescence Staining

Journal: Science translational medicine

Article Title: Hematopoietic loss of Y chromosome activates immune checkpoints and contributes to impaired senescent cell clearance and renal disease

doi: 10.1126/scitranslmed.adv4071

Figure Lengend Snippet: ( A ) Flow cytometric detection of immune inhibitory receptors (left, PD-1; right, SIRPα) on the indicated immune cell types in kidneys of aged LOY and CT mice at 15 months after BMT (CT: n = 3; LOY: n = 3). Statistical significance was determined by a two-way ANOVA with Tukey’s post hoc test. ( B ) Flow cytometric analyses for PD-1 and SIRPα immune inhibitory receptor expression on the indicated immune cell types in injured kidneys of LOY and CT mice at 4 weeks after the administration of AA (CT: n = 6; LOY: n = 5). Statistical significance was determined by unpaired Student’s t tests. ( C ) Schematic of experiment assessing kidney injury in the hematopoietic LOY model. A series of peritoneal injections every 3 days with combined PD-1 and SIRPα ICI antibodies in LOY and CT mice was started 1 week after the administration of AA, and renal phenotypes were evaluated 4 weeks after the administration of AA. ( D ) Serum BUN amounts (CT control: n = 6; LOY control: n = 5; CT ICIs: n = 6; LOY ICIs: n = 6). Statistical significance was determined by a one-way ANOVA with Tukey’s post hoc test. ( E ) qRT-PCR analysis of kidney injury marker genes in kidneys (CT control: n = 6; LOY control: n = 5; CT ICIs: n = 6; LOY ICIs: n = 6). Statistical significance was determined by unpaired Student’s t tests. ( F ) qRT-PCR analysis of senescence-associated genes in kidneys (CT control: n = 6; LOY control: n = 5; CT ICIs: n = 6; LOY ICIs: n = 6). Statistical significance was determined by a one-way ANOVA with Tukey’s post hoc test. ( G ) Representative images and quantitative analysis of senescent area in kidneys, as stained with SA-β-gal (CT control: n = 6; LOY control: n = 5; CT ICIs: n = 6; LOY ICIs: n = 6). Scale bars, 50 μm. Statistical significance was determined by unpaired Student’s t tests. ( H ) qRT-PCR analysis of the fibrosis-related gene Acta2 in kidneys (CT control: n = 6; LOY control: n = 5; CT ICIs: n = 6; LOY ICIs: n = 6). Statistical significance was determined by a one-way ANOVA with Tukey’s post hoc test. ( I ) Representative images and quantitative analysis of fibrotic area in kidneys, as stained with picrosirius red (CT control: n = 6; LOY control: n = 5; CT ICIs: n = 6; LOY ICIs: n = 6). Scale bars, 50 μm. Statistical significance was determined by unpaired Student’s t tests. Dots in all panels represent individual samples. Data are shown as the mean ± SEM (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, and NS ≥ 0.05).

Article Snippet: To evaluate the effect of treatment with ICIs on LOY-mediated renal damage in the murine model of AA-induced fibrosis and senescence, an anti–PD-1 antibody (Bio X Cell, no. BE0146) and an anti-SIRPα antibody (Bio X Cell, no. BE0322) were administered to hematopoietic mosaic LOY and CT mice 1 week after the administration of AA.

Techniques: Expressing, Control, Quantitative RT-PCR, Marker, Staining

Journal: bioRxiv

Article Title: Binding of inhibitory checkpoints to CD18 in cis hinders anti-cancer immune responses

doi: 10.1101/2025.09.10.675342

Figure Lengend Snippet: ( A ) Existing model by which SIRPα suppresses phagocytosis by interacting in trans with CD47 on target cells. See text for details. The 3 Ig-like domains of SIRPα (1 IgV and 2 IgCs) and the single Ig-V domain of CD47 are shown as ellipses. Mβs, macrophages. ( B ) Depiction of SIRPα variants and their functional characteristics. SIRPα FFFF contained substitution of tyrosine (Y)-to-phenylalanine (F) substitution at Y436, 460, 477, and 501; SIRPα ΔIC lacked most of the cytoplasmic domain of SIRPα, ending with arginine 401; SIRPα T96V carried a threonine (T)-to-valine (V) mutation at position 96 (shown by lavender star), which abolishes CD47-binding; SIRPα T96V,FFFF had the T96V and FFFF mutations; SIRPα T96V,ΔIC had the T96V and the ΔIC mutations. KO, knock-out. ITIM, immunoreceptor tyrosine-based inhibitory motif. ( C to G ) SIRPα variants or empty vector were expressed in SIRPα KO BMDMs and tested. Wild-type (WT) BMDMs were used as control. ( C ) Schematic representation of assays performed. Fc, fragment crystallizable. ( D ) Flow cytometry analyses of SIRPα expression and CD47-binding. APC, allophycocyanin. AF647, Alexa fluor 647. ( E and F ) Representative ( E ) and compiled data ( F ) of pHrodo-based phagocytosis assays using L1210 derivatives expressing Tac and opsonized with Tac monoclonal antibody (mAb) 7G7, as targets. Positive cells with percentages are boxed. G , Efficiency of phagocytosis inhibition in SIRPα KO BMDMs expressing or not the indicated SIRPα variants was calculated using the values in ( F ). SIRPα KO expressing WT SIRPα or empty vector displayed 100% and 0% inhibition efficiency, respectively. All data are means ± s.e.m., **** p < 0.0001. Results in ( D and E ) are representative of 6 independent experiments, except for SIRPα T96V , SIRPα T96V, FFFF and SIRPα T96V, ΔIC that are representative of 3 experiments. Results in ( F and G ) are pooled from a total of 6 mice studied in 6 independent experiments, except for SIRPα T96V , SIRPα T96V, FFFF and SIRPα T96V, ΔIC that involved 3 mice in 3 experiments. Each symbol in ( F ) represents one mouse.

Article Snippet: Human SIRPα V2 cDNA (Cat: SC125649) was from OriGene (Rockville, MD).

Techniques: Functional Assay, Mutagenesis, Binding Assay, Knock-Out, Plasmid Preparation, Control, Flow Cytometry, Expressing, Inhibition

Journal: bioRxiv

Article Title: Binding of inhibitory checkpoints to CD18 in cis hinders anti-cancer immune responses

doi: 10.1101/2025.09.10.675342

Figure Lengend Snippet: ( A and B ) Immunoprecipitation followed by mass spectrometry of SIRPα-associated proteins. ( A ) Schematic representation of assay. ( B ) Plasma membrane-associated proteins found in SIRPα immunoprecipitates from WT BMDMs, but not from SIRPα KO BMDMs. c , Co-immunoprecipitation assay of SIRPα, CD18 and CD11b in WT and SIRPα KO BMDMs. IP, immunoprecipitation. Abs, antibodies. ( D to F ) FRET assays. ( D ) Schematic representation of FRET assay in HEK293T cells. ( E and F ) Representative confocal microscopy images ( E ) and compiled data ( F ) of FRET assays with donor-labeled SIRPα, acceptor-labeled CD18 and unlabeled CD11b in the presence of control (Ctrl) IgG, CD18 mAb GAME-46 or CD11b mAb 5C6. Yellow to purple spectrum denotes strong to weak FRET. DIC, differential interference contrast. Scale bars, 5 μm. ( G and H ) LUV-FRET assay. ( G ) Schematic representation of LUV-FRET assay. ( H ), Time-course of donor-labeled SIRPα fluorescence intensity after addition of acceptor-labeled CD18 or CD11b, monitored with a real-time plate reader. All data are means ± s.e.m. ns, not significant, **** p < 0.0001. Results in ( C , E and H ) are representative of 3 independent experiments. Results in ( B and F ) are pooled from a total of 3 independent experiments. Each symbol in ( F ) represents one cell.

Article Snippet: Human SIRPα V2 cDNA (Cat: SC125649) was from OriGene (Rockville, MD).

Techniques: Immunoprecipitation, Mass Spectrometry, Clinical Proteomics, Membrane, Co-Immunoprecipitation Assay, Confocal Microscopy, Labeling, Control, Fluorescence

Journal: bioRxiv

Article Title: Binding of inhibitory checkpoints to CD18 in cis hinders anti-cancer immune responses

doi: 10.1101/2025.09.10.675342

Figure Lengend Snippet: ( A and B ) FRET assays with SIRPα and SIRPβ1a. ( A ) A schematic representation of SIRPα and SIRPβ1a, with their 1 IgV domain and 2 IgC domains, is depicted. ( B ) Compiled data of 3 independent experiments using donor-labeled SIRPα or SIRPβ1a, acceptor-labeled CD18 and unlabeled CD11b, as done for , D to F. ( C and D ) FRET assays using SIRPα IgV domain. ( C ) A schematic representation of a SIRPα variant having only the IgV domain is shown. ( D ) Compiled data of 3 independent experiments using donor-labeled SIRPα IgV, acceptor-labeled CD18 and unlabeled CD11b, as done for , D to F. ( E - H ) FRET assays using SIRPα variants carrying non-conserved residues from SIRPβ1a. ( E and G ) Schematic representations of SIRPα variants. ( F and H ) Compiled data from 3 independent experiments using donor-labeled SIRPα variants, acceptor-labeled CD18 and unlabeled CD11b, as done for , D to F. ( I to K ) Proximity ligation assay (PLA) of SIRPα and CD18 in BMDMs expressing or not the indicated SIRPα variants. (I) Flow cytometry analyses of SIRPα expression. ( J and K ) Representative confocal microscopy images ( J ) and compiled data from 3 independent experiments ( K ) of PLA for SIRPα and CD18. Scale bar, 10 μm. All data are means ± s.e.m. ns, not significant, **** p < 0.0001. Results in ( I and J ) are representative of 3 independent experiments. Results in ( B , D , F , H and K ) are pooled from 3 independent experiments. Each symbol in ( B , D , F , H and K ) represents one cell or mouse.

Article Snippet: Human SIRPα V2 cDNA (Cat: SC125649) was from OriGene (Rockville, MD).

Techniques: Labeling, Variant Assay, Proximity Ligation Assay, Expressing, Flow Cytometry, Confocal Microscopy

Journal: bioRxiv

Article Title: Binding of inhibitory checkpoints to CD18 in cis hinders anti-cancer immune responses

doi: 10.1101/2025.09.10.675342

Figure Lengend Snippet: ( A to C ) The impact of SIRPα variants defective in CD18-binding, CD47-binding or phosphatase signaling, alone or in combination, expressed in BMDMs, was analyzed. ( A ) Schematic depictions of SIRPα variants, as was done for . SIRPα R91T carried an arginine (R)-to-threonine (T) mutation at position 91 (shown by blue star), which abolished CD18-binding. ( B ) Phagocytosis assays of IgG-opsonized L1210 cells by BMDMs, as was done for . ( C ) Efficiency of phagocytosis inhibition was calculated as for , using values from . ( D and E ) Representative flow cytometry profiles ( D ) and compiled data from 3 independent experiments ( E ) of ICAM-1-binding using SIRPα KO BMDMs expressing WT SIRPα or SIRPα R91T BMDMs, in the presence or absence of FcR triggering using mouse IgG2a. ( F and G ) The impact of a SIRPα variant carrying the isoleucine-to-glycine 332 (I332G) mutation, expressed in SIRPα KO BMDMs, was analyzed. (F) Flow cytometry analyses of CD11b expression. ( G ) Compiled data from 3 independent phagocytosis assays, assessed by microscopy. ( H ) FRET assays of donor-labeled SIRPα, acceptor-labeled CD18 and unlabeled CD11b in the presence of WT CD11b or CD11b I332G , as was done for , D to F. ( I ) FRET assays of donor-labeled human SIRPα version (V) 1 or V2 with acceptor-labeled human CD18 and unlabeled human CD11b, in the presence of Ctrl IgG, human CD18 mAbs CBR LFA1/2 or TS1/18, as was done for , D to F. ( J ) Phagocytosis of human lymphoma cells Raji, which were opsonized with CD20 mAbs, by human peripheral blood monocyte (PBMC)-derived macrophages, in the presence of the indicated mAbs, was assessed by microscopy. All data are means ± s.e.m. ns, not significant; * p < 0.05, ** p < 0.01 and **** p < 0.0001. Results in ( D and F ) are representative of 3 independent experiments. Results in ( B , C , E and G to J ) are pooled from 3 independent experiments. Each symbol in ( B , E and G to J ) represents one cell, mouse or healthy donor.

Article Snippet: Human SIRPα V2 cDNA (Cat: SC125649) was from OriGene (Rockville, MD).

Techniques: Binding Assay, Mutagenesis, Inhibition, Flow Cytometry, Expressing, Variant Assay, Microscopy, Labeling, Derivative Assay

Journal: bioRxiv

Article Title: Binding of inhibitory checkpoints to CD18 in cis hinders anti-cancer immune responses

doi: 10.1101/2025.09.10.675342

Figure Lengend Snippet: ( A ) FRET assays of donor-labeled mouse SIRPα with acceptor-labeled mouse CD18 and unlabeled mouse CD11b, in the presence of Fc-silent mouse SIRPα mAbs, as was done for , D to F. ( B ) Binding of a soluble CD47-Fc fusion protein to EL-4 cells, expressing or not expressing mouse SIRPα, was studied by flow cytometry. ( C to K ) Generation and impact of bispecific antibody (BsAb) against mouse SIRPα. ( C ) Schematic representation of Fc-silent BsAb combining one arm of mAb #17 with one arm of mAb #27, using the “knob-into-hole” technology. Phagocytosis of IgG-opsonized L1210 cells ( D ) and EL-4 cells ( E ) by WT BMDMs, in the presence of mAbs, was assessed by a microscopy assays. ( F to K ) Schematic depictions of the assays are shown in (F and I). RAG-1 KO mice injected subcutaneously with Tac + L1210 cells ( G and H ), or C57BL/6J mice injected subcutaneously with Tac + EL-4 cells ( J and K ), were treated by intraperitoneal injection of Fc-silent mAbs, alongside Tac mAb 7G7 for opsonization. Tumor volume was measured using a caliper ( G and J ) and survival was recorded ( H and K ). ( L ) FRET assays of donor-labeled human SIRPα V1 or V2 with acceptor-labeled human CD18 and unlabeled human CD11b in the presence of Fc-silent Ctrl IgG and human SIRPα mAbs KWAR23, 40A, 50A, or 18D5, as was done for , D to F. The mAbs were rendered Fc-silent by the LALAPG mutation. ( M ) Phagocytosis of IgG-opsonized Raji cells by human macrophages in the presence of Fc-silent Ctrl IgG and SIRPα mAbs KWAR23, 40A, 50A, or 18D5, was assayed as for . ( N ) FRET assays of donor-labeled human 2B4 (SLAMF4), PD-1 or LILRB1 with acceptor-labeled human CD18, in the presence of Ctrl IgG or human CD18 mAb were done as for , D to F. All data are means ± s.e.m. ns, not significant; * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001. Results are pooled from a total of two ( H and K ), three ( A , D , E , G , J , L and N ) or five ( B and M ) independent experiments. Each symbol in ( A , D , E and L to N ) represents one healthy donor, cell or mouse.

Article Snippet: Human SIRPα V2 cDNA (Cat: SC125649) was from OriGene (Rockville, MD).

Techniques: Labeling, Binding Assay, Expressing, Flow Cytometry, Microscopy, Injection, Mutagenesis