53-0869-41

antibody from Invitrogen Antibodies

Targeting: CD86 B7-2, B7.2, CD28LG2

Provider product page for 53-0869-41

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Antibody data

Antibody Data
Antigen structure
References [16]
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Validations
Flow cytometry [1]
Other assay [18]

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Product number: 53-0869-41 - Provider product page
Provider: Invitrogen Antibodies
Product name: CD86 (B7-2) Monoclonal Antibody (IT2.2), Alexa Fluor™ 488, eBioscience™
Antibody type: Monoclonal
Antigen: Other
Description: Description: The IT2.2 monoclonal antibody reacts with human CD86, an ~80 kDa surface receptor also known as B7-2. CD86 and CD80 are members of the B7 family of costimulatory molecules. CD86 is expressed at low levels on B cells, macrophages, and dendritic cells and is upregulated on B cells through a variety of surface stimuli including the BCR complex, CD40 and some cytokine receptors. In addition to CD80 (B7-1), CD86 is a counter-receptor for the T cell surface molecules CD28 and CD152 (CTLA-4). The interaction of CD86 with its ligands plays a critical role in T-B crosstalk, T cell costimulation, autoantibody production and Th2-mediated Ig production. The kinetics of upregulation of CD86 upon stimulation supports its major contribution during the primary phase of an immune response. Applications Reported: This IT2.2 antibody has been reported for use in flow cytometric analysis. Applications Tested: This IT2.2 antibody has been pre-titrated and tested by flow cytometric analysi sof normal human perioheral blood cells. This can be used at 5 µL (0.25 µg) per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. Excitation: 488 nm; Emission: 519 nm; Laser: Blue Laser. Filtration: 0.2 µm post-manufacturing filtered.
Reactivity: Human
Host: Mouse
Conjugate: Green dye
Isotype: IgG
Antibody clone number: IT2.2
Vial size: 25 Tests
Concentration: 5 µL/Test
Storage: 4° C, store in dark, DO NOT FREEZE!

CD86 protein structure - 53-0869-41 shown in red.

Supportive validation

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Experimental details: NULL
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 miR-720 suppresses the expression of M2 phenotypes ( A ) Real-time PCR analysis of miR-720 expression in control vector or pLV- miR-720 -infected THP-1 cells. ( B ) Control or pLV- miR-720 -infected THP-1 cells were induced macrophage differentiation by PMA (M0) and then treated with IFN-gamma plus LPS (M1) or IL-4 plus IL-13 (M2) to induce polarization. The macrophage marker CD68, M1 marker CD86 and M2 marker CD163 were analysed by flow cytometry respectively. ( C ) Detection of M1 cytokine TNF-alpha and IL-6 or ( D ) M2 cytokine IL-10 and chemokine CCL17 by real-time PCR in THP-1 cells used in ( B ). Values are means +- S.D. ** P
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 4 Silencing PTX3 in apoptotic cells raised the affinity for macrophages and attenuated inflammatory responses. (A) Binding of apoptotic bodies on phagocytes was elevated in PTX3(-) THP-1 cells (means+-SD, n =8, * p
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 5 DCs maturation, T cell proliferation, and IDO expression in the co-culture system. (A) Flow cytometry detection of CD86 and MHC-II DC surface markers; mean fluorescence intensity (MFI) was calculated at 72 hpi. CD86 and MHC-II were analyzed by pre-gated CD11c + cells, CD11c was gated by isotype. CD86 and MHC-II surface markers were significantly upregulated in the co-culture system of CD4 + T cells and IL-10 -/- DCs. Compared to the DCs from the zDC-DTR group, CD86 and MHC-II were significantly higher in the DD group. (B) Measurement of CD4 + T cell proliferation via BrdU assay in the co-culture system of CD4 + T cells and DCs at 72 h. Higher CD4 + T cell proliferation was observed in the IL-10 -/- and anti-IL-10 groups than the zDC-DTR group ( P
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 3. Treatment with RB-3 leads to reduction in H2Aub and differentiation in TEX cells. a , Assessment of H2Aub, H2Bub, H3 and total H2A in TEX cells treated with RB-3 (left) or RB-nc (right) for 4 days. Representative gel of three independent experiments. b , Effect of RING1B inhibition on the growth of TEX cells treated with RB-3 and RB-nc. Representative data from two independent experiments. c, d, e, Flow-cytometry analysis of CD34 and CD38 ( c ), CD11b ( d ) and CD86 ( e ) expression in TEX cells treated with 25 muM RB-3 and 25 muM RB-nc for 21 days. Representative histograms of two independent experiments. f, Cell morphology of TEX cells after 21 days of treatment with 25 muM RB-3, 25 muM RB-nc or DMSO analyzed by Wright Giemsa staining. Representative slides are shown from two independent experiments.
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 6. RB-3 reduces global H2Aub levels and induces differentiation in primary AML patient samples. a, Western blot detection of H2Aub, and H3 in four primary AML patient samples treated with RB-3 for 12 days. Western blots were performed once due to limited amounts of primary samples. b , Colony counts from the second round of colony forming assay in primary AML patient samples treated with 25 muM RB-3 for 14 days. Data are two technical replicates from single experiment. c , Quantification of CD34 + cells in primary AML patient samples treated with 25 muM RB-3 for 12 days. Data are two technical replicates from single experiment. d, Representative FACS histograms showing the level of CD34 + cells for sample #3 treated with 25 muM RB-3 and 25 muM RB-nc for 12 days. e , Flow cytometry quantitation of CD11b + cells in primary AML samples treated with 25 muM RB-3 for 12 days. Data are two technical replicates from single experiment. f , Representative FACS histograms showing the level of CD11b + cells for sample #1 treated with 25 muM RB-3 and 25 muM RB-nc. g, Western blot detection of C/EBPalpha-p42 and beta-actin levels in AML samples treated with RB-3 for 12 days. Western blots were performed once. h , Representative FACS histograms showing CD86 level in sample #4 treated with 25 muM RB-3 and 25 muM RB-nc for 12 days. i , Cell morphology of AML cells analyzed by Wright-Giemsa Giemsa staining after 14 days treatment with 25 muM RB-3 or DMSO. Representative slides from sing
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Extended Data Fig. 6 Long-term effect of treatment of TEX cells with RB-3. a ) Western blot detection of H2Aub in TEX cells treated with increasing doses of RB-3 and RB-nc for 21 days. Representative blot out of two replicates. b ) Western blot detection of total cellular ubiquitination levels using ubiquitin specific antibody in TEX cells treated with RB-3 for 21 days. Representative blot out of three replicates. c ) Flow cytometry analysis of CD34 in TEX cells treated with RB-3 for 7 days. On day 7, cells were stained with APC/CY7-conjugated human anti-CD34 antibody and analyzed by FACS. Representative histograms of two independent experiments. d ) Flow cytometry analysis of CD34 and CD38 in TEX cells treated with RB-3 (top panel) and RB-nc (bottom panel) for 21 days. On day 21, cells were stained with APC/CY7-conjugated human CD34 and PE-conjugated CD38 antibodies and analyzed by FACS. Representative histograms of two independent experiments. e,f ) Flow cytometry analysis of myeloid differentiation marker CD11b/ ITGAM in TEX cells treated with RB-3 for 7 days ( e ) and with RB-3 ( f , top panel) and RB-nc ( f , bottom panel) for 21 days. Cells were stained with Pacific Blue human CD11b antibody and analyzed by FACS. Representative histograms of two independent experiments. g ) Flow cytometry analysis of dendritic cells differentiation marker CD86/B7-2 in TEX cells treated with RB-3 (top panel) and RB-nc (bottom panel) for 21 days. On day 21, cells were stained with Super B
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: The correlation between TAMs and HNSCC prognosis and the establishment of HOTTIP overexpression and knockdown exosomes in M1. A The infiltration levels of immune cells in HNSCC. B Prediction of overall survival according to high- and low-infiltration levels of M0, M1 and M2 macrophages. THP-1-derived M0, M1 and M2 macrophages were confirmed by flow cytometry using ( C ) CD14 plus CD86 and ( D ) CD14 plus CD163 ( n = 3). E The exosomes were extracted from three types of macrophages and confirmed by electronic microscopy and Western blot analysis using CD9 and CD63. F RT-qPCR was conducted to test the expression of HOTTIP in M0, M1 and M2 macrophages ( n = 3). RT-qPCR was conducted to test the expression of HOTTIP in wild type M1 macrophages, HOTTIP-overexpressed and HOTTIP-knockdown M1 macrophages ( G ) and their exosomes ( H ) ( n = 3). Data are presented as mean +- SD. Results were analyzed by One-way ANOVA with a post hoc t -test. Significance: * P < 0.05, ** P < 0.01, *** P < 0.001.
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Cancer cells expressing HOTTIP and M1 exosomes reprogram the circulating monocytes into M1 phenotype. A-D Flow cytometry assay detected CD14 + CD86 + M1 and CD14 + CD163 + M2 phenotype in circulating blood of nude mice bearing with HOTTIP-knockdown Hep-2 cells ( n = 3). HOTTIP-knockdown Hep-2 cells suppressed the polarization of M1 monocytes ( A , C ) but induced the polarization of M2 ( B , D ). E - H Flow cytometry assay detected M1 and M2 phenotypes in the blood of tumor-bearing nude mice treated by M1 exosomes ( n = 3). M1 exosomes induced the polarization of M1 monocytes ( E , G ) but inhibited the polarization of M2 ( F , H ). Data are presented as mean +- SD. Results were analyzed by One-way ANOVA with a post hoc test. Significance: ns not significant, ** P < 0.01, *** P < 0.001.
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 CFT073 wt infection significantly inhibits CD86 and iNOS but promotes CD206 and Arg-1 expression in K-macrophages from PN mouse models. ( A , B ) Flow cytometry analyses to examine the expression of CD86 and CD206 on K-macrophages. ( C ) FI of CD86 and CD206 were analyzed by FlowJo v10.0.7 software. FI in the control group was set as 1.0. ( D ) mRNA levels of iNOS and Arg-1 in K-macrophages were detected by qRT-PCR. ( E ) Protein levels of iNOS and Arg-1 in K-macrophages were detected by Western blot. ( F ) Gray-scale analyses of bands reflecting iNOS and Arg-1 protein levels in experiments as described in E. Gray scale in the control group was set as 1.0, Mean +- SD of three independent experiments are shown. *: p < 0.05; **: p < 0.01.
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 3 CFT073 wt suppresses M1 but promotes M2 macrophage polarization in vitro. ( A , B ) The expression of M1 surface markers CD80 and CD86 in THP-1 and J774A.1 was detected by flow cytometry. ( C , D ) The expression of M2 surface markers CD163 and CD206 in THP-1 and J774A.1 was analyzed by flow cytometry. ( E ) The production of NO in THP-1 and J774A.1 was measured by Griess method. ( F , G ) The iNOS protein levels and bands' gray-scale analyses in THP-1 and J774A.1, respectively. ( H ) The protein levels of Arg-1 in THP-1 and J774A.1 were detected by ELISA. ( I , J ) The protein levels of Arg-1 were detected by Western blot and bands' gray-scale analyses in THP-1 and J774A.1, respectively. Gray scale in the control group was set as 1.0. Mean +- SD of three independent experiments are shown. *: p < 0.05; **: p < 0.01. NS: not significant.
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: FIGURE 2 Flow cytometry analysis of M1 and M2 surface markers (A) The expression of CD86 (M1 surface marker) and CD206 (M2 surface marker) on RAW264.7 cells were examined by flow cytometry. (B) Quantification of mean fluorescence intensity (MFI) of the surface markers ( n = 3, respectively). (C) The expression of CD86 and CD206 on THP-1 cells were examined by flow cytometry. (D) Quantification of mean fluorescence intensity (MFI) of the surface markers ( n = 3, respectively). The results are expressed as the mean values +- S.E.M. * p < 0.05, ** p < 0.01, *** p < 0.001.
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2. Percentage of purified DC samples in all experiments. DCs were labeled with the Non-DC Depletion Cocktail for negative selection. Next, DCs were labeled with DC Enrichment Cocktail for positive selection. The expression of CD11c and CD86 was tested by flow cytometery. The percentages of purified DC are presented in the top of each panel. DCs, dendritic cells; CD, cluster of differentiation.
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 1. The remnants of LM8 decreased the co-stimulatory molecules, and inhibited IL-12 and IL-6 levels, increased IL-10 levels, and inhibited the ability of DCs to stimulate T-cell proliferation. (A) The DCs were isolated following a 48-h co-culture with the remnants of LM8 cells. The cells were then labeled with antibodies against CD11c, MHC-2, CD40, CD86, CD80 and CCR7 for phenotypic analysis by flow cytometry. The numbers in the histograms indicate the geometric mean fluorescence intensity. (B) Following isolation from the co-culture system, the DCs were cultured for 12 h. Subsequently, the expression levels of IL-12, IL-6 and IL-10 in the supernatant were analyzed by ELISA. (C) CD4 + T cells from DO11.10 OVA 323-339 -specific (TCR-transgenic x C57BL/6) F1 hybrid mice were co-cultured with DCs or mDCs (control) in the presence of OVA peptides. Five days later, the total number of viable CD4 + T (CD4 + 7-AAD - ) cells in each well was measured by flow cytometry. Data represent one of at least three experiments with similar results. *P
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2. PDT treatment partly reversed the effect of LM8 remnants on the phenotype of DCs and their ability to stimulate T cells proliferation. (A) The LM8 cells were pre-treated with PDT, and then labeled with antibodies against MHC-2, CD11c, CD40, CD86, CD80 and CCR7, for phenotypic analysis by flow cytometry. (B) The numbers in the histograms indicate the geometric mean fluorescence intensity. *P
Conjugate: Green dye

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 3. PDT treatment upregulated the HSP70 expression in LM8 cells and promoted upregulation of HSP70-activated DCs. (A) The LM8 cells with and without PDT pre-treatment were collected for RNA sequencing analysis. (B) The expression of HSP70, ATF3, Bcl-2, P53, P21, P16 and P27 was analyzed by reverse transcription-quantitative PCR. (C) LM8 cells were transfected with HSP70 small interfering RNA, and the DCs were then co-cultured with LM8 for 48 h, prior to being labeled with an antibody against CD86 for phenotypic analysis by flow cytometry. The numbers in the histograms indicate the geometric mean fluorescence intensity. Data represent one of at least three experiments with similar results. *P
Conjugate: Green dye