PP was provided by the Kangmei Pharmaceutical Co. Ltd. (Guangdong, China).PEGDA (containing 400–600 ppm MEHQ stabilizer) was purchased from the Shanghai Yuanye Biotechnology Co. Ltd. (China). CMC was purchased from the Zhejiang Aoxing Biotechnology Co. Ltd. (China). GO was obtained from the Xianfeng Nanomaterials Technology Co. Ltd. (Jiangsu, China). 2-Hydroxy-4-(2-hydroxyethoxy)-2-methylpropiophenone (Photoinitiator 2959) was purchased from Aladdin Reagent (Shanghai, China). Interleukin (IL)-6, IL-4, IL-10, transforming growth factor-β (TGF-β), and tumor necrosis factor-α (TNF-α) ELISA kits were purchased from YIFEIXUE BIOTECH (Nanjing, China). FITC-conjugated anti-CD86, PE-conjugated anti-CD206, recombinant anti-iNOS, anti-osteocalcin (OCN), and anti-alkaline phosphatase (ALP) antibodies were purchased from Thermo Fisher Scientific (Waltham, MA, USA). CD68 rabbit polyclonal, bone morphogenetic protein-2 (BMP-2) polyclonal, runt-related transcription factor 2 (RUNX2), and osteopontin (OPN) polyclonal antibodies were purchased from AiFang Biological (Hunan, China). Osteogenic inducting fluid was purchased from Procell (Wuhan, China). ALP Color Development Kit and Alizarin red S (ARS) kit were purchased from Beyotime (Shanghai China).
Fitc conjugated anti cd86
Manufactured by Thermo Fisher Scientific
Sourced in United States
FITC-conjugated anti-CD86 is a primary antibody that binds to the CD86 antigen. CD86 is a costimulatory molecule expressed on the surface of antigen-presenting cells. The FITC (fluorescein isothiocyanate) label allows for the detection and analysis of CD86-positive cells using flow cytometry or fluorescence microscopy.
Automatically generated - may contain errors
Lab products found in correlation
Alp color development kit, by Beyotime (1 mentions)
Facsverse, by BD (1 mentions)
Anti pd l1, by Thermo Fisher Scientific (1 mentions)
Mouse regulatory t cell staining kit, by Thermo Fisher Scientific (1 mentions)
Fitc conjugated anti mouse cd4, by Thermo Fisher Scientific (1 mentions)
Apc conjugated anti mouse cd3, by Thermo Fisher Scientific (1 mentions)
Apc conjugated anti mouse cd11b, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti mouse cd8, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti mouse gr 1, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti mouse pd 1, by BD (1 mentions)
Apc conjugated anti cd11c, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti mhc 2, by Thermo Fisher Scientific (1 mentions)
Mouse seroblock fcr, by Bio-Rad (1 mentions)
E coli o55 b5, by Merck Group (1 mentions)
Facscanto 2 cytometer, by BD (1 mentions)
Fitc conjugated anti cd11b, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti f4 80, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti cd80, by Thermo Fisher Scientific (1 mentions)
Facscalibur flow cytometer, by BD (1 mentions)
5 protocols using fitc conjugated anti cd86
1
Polymer-Based Tissue Engineering Scaffold
2
Flow Cytometry Analysis of Macrophage Activation
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We used murine FITC-conjugated anti-CD86 or murine PE-conjugated anti-MHCII antibodies from Thermo Fisher (Waltham, MA, USA) to stain the activation markers CD86 and MHCII, respectively. RAW 264.7 was incubated with the antibodies for 30 minutes at 4°C in FACS buffer (PBS supplemented with 4% FBS) and then washed with FACS buffer again. The cells were also fixed with paraformaldehyde 1% for 5 minutes and permeabilized with Triton 0.1% for 5 minutes for iNOS labeling. Then, cells were incubated with murine Alexa Fluor 488-conjugated antibody for 30 minutes at 4 ºC in FACS buffer and washed with FACS buffer again. We used flow cytometry (model FACSVerse, BD Biosciences, San Jose, USA) for data acquisition and the FlowJo software (San Jose, CA, USA) for the analysis of the median fluorescence intensity values (MFI) of iNOS, MHCII, and CD86 expression, which were normalized by the mean of negative control MFI. The percentage of double-labeled cells was used to show the population of cells expressing MHCII and CD86 simultaneously.
3
Comprehensive Immunophenotyping of T-cells and DCs
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T-cell suspensions were isolated from the spleen and lymph nodes, and were stained with antibodies against the following cell surface antigens: APC-conjugated anti-mouse CD3, FITC-conjugated anti-mouse CD4, PE-conjugated anti-mouse CD8, PE-conjugated anti-mouse Gr-1, APC-conjugated anti-mouse CD11b, PE-conjugated anti-mouse TCR (eBioscience), PE-conjugated anti-mouse PD-1, and PE-conjugated anti-mouse CD152 (CTLA-4; BD Bioscience, San Jose, CA, USA). Tregs were detected from the spleen and lymph nodes using a Mouse Regulatory T-Cell Staining Kit (eBioscience) according to the manufacturer's instructions. DCs were isolated from the bone marrow and cells were incubated with FITC-conjugated anti-CD86, APC-conjugated anti-CD11C, PE-conjugated anti-MHC-II, FITC-dextran, PE-conjugated anti-CCR7, and anti-PD-L1 (eBioscience). The cells were analyzed by FACS, and the acquired data was performed with FlowJo Software, version 9.1 (Tree Star, San Carlos, CA, USA).
4
Immunological Response to Klebsiella pneumoniae
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Groups of 8-wk-old male BALB/c mice were intraperitoneally injected with 200 ul PBS containing with and without LPS (1 mg/Kg of weight; E. coli O55:B5; Sigma). After 16 hours, the LPS-stimulated mice were intraperitoneally challenged with 1 × 109 CFU of K. pneumoniae KPC160111 or KPC160132. Peritoneal cells were harvested by injecting 5 mL of PBS at 1.5 h post-bacterial challenge, and the supernatants were collected for cytokine/chemokine analysis. After RBC lysis and washing, peritoneal cells were treated with Mouse Seroblock FcR (Bio-Rad Laboratories # BUF041) for 20 min at 4°C to block the Fc receptors and then incubated with rat anti-mouse fluorescent antibodies to comparatively analyze the distribution of the myeloid cells in response to K. pneumoniae challenge. The antibodies used were PE-conjugated anti-F4/80 (eBioscience #12-4801-82), FITC-conjugated anti-CD11b (eBioscience #11-0112-82), FITC-conjugated anti-CD86 (eBioscience #11-0862-82), and PE-eFluor 610-conjugated anti-CD163 (eBioscience #61-1631-82). After incubation at 4°C for 1 h, cells were washed and analyzed with a BD FACS Canto II cytometer (Becton Dickinson). Flow cytometry data were analyzed with FlowJO v10.7 (Becton Dickinson).
5
Measuring Immune Cell Activation Markers
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Antigen-presentation markers (MHC class I and II) and co-stimulation markers (CD80 and CD86) were measured in BMMs by flow cytometry after treatment with LPS (50 ng/ml) or TgPLP (500 ng/ml) for 24 hours. Antibodies for immunostaining were as follows: APC-conjugated anti-MHC1 (1:100, eBioscience, CA), APC-conjugated anti-MHC2 (1:100, eBioscience), PE-conjugated anti-CD80 (1:100, eBioscience), FITC-conjugated anti-CD86 (1:100, eBioscience). For immunostaining, BMMs were incubated with each antibody for 30 minutes at 4°C, washed with FACS buffer three times, fixed in 4% paraformaldehyde, and analyzed by FACSCalibur flow cytometer (BD science). MFI was measured by a FlowJo FACS analyzer (Tree Star).
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