The largest database of trusted experimental protocols

41 protocols using anti cd11b fitc

1

Multiparameter Analysis of Hematopoietic Stem Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols
The following commercial antibodies were used: mouse hematopoietic lineage eFlour 450 cocktail, PerCP-Cy5.5-anti-CD45.1, FITC-anti-CD45.2, Alexa Fluor 700-anti-IL-7Rα, FITC-anti-Ly6A/E (Sca-1), PE-anti-CD117 (c-Kit), APC-eFluor780-anti-CD48, anti-CD3, anti-CD19, FITC-anti-CD11b, PE-anti-Gr1, FITC-anti-B220, Pecy5-anti-CD3ε, APC-anti-Flt3/CD135, anti-F4/80, anti-Gr-1, and anti-CD34 were purchased from eBioscience. PE-cy7-anti-CD150 was obtained from BioLegend. PerCP-Cy5.5–conjugated goat anti–rat IgG and APC-Cy7–conjugated goat anti–rabbit IgG were purchased from Santa Cruz Biotechnology, Inc. Anti–β-actin was from Sigma-Aldrich. Antibodies against Insr-β, phosphorylated Insr-β (Tyr1150/1151), insulin, mTOR, Stat3, S727 phosphorylated Stat3, Y705 phosphorylated Stat3, S6K, and phosphorylated S6K were purchased from Cell Signaling Technology. Donkey anti–rabbit or anti–mouse secondary antibodies conjugated with Alexa Fluor 488, 594, or 405 were purchased from Molecular Probes. HRP-conjugated secondary antibody was obtained from Santa Cruz Biotechnology, Inc. Propidium iodide (PI), Annexin-V, insulin, rapamycin, and streptozotocin (STZ) were purchased from Sigma-Aldrich.
+ Open protocol
+ Expand
2

Oroxylin A Purification and Inflammatory Assays

Check if the same lab product or an alternative is used in the 5 most similar protocols
Oroxylin A was isolated from the root of Scutellaria baicalensis as previously described [31 (link)]. Samples containing oroxylin A at a minimum of 99% purity were used for the experiments unless otherwise indicated. LPS, PMA and ATP were purchased from Sigma Aldrich (St. Louis, MO, USA). DSS (molecular weight 36–50 kDa) was obtained from MP Biomedicals Inc. (Irvine, CA, USA). Antibodies against p-IκBα, IκBα and p65 were purchased from Cell Signaling Technology (Danvers, MA, USA). Primary antibodies against lamin A, β-actin, IL-1β and bay11-7082 were purchased from Bioworld Technology Inc. (CA, USA). Exfect Transfection reagent was purchased from Vazyme (Nanjing, China). NLRP3 and caspase-1 antibodies were from Abcam Technology Inc (MA, USA). FITC-anti-CD11b and FITC-anti-F4/80+ were purchased from eBioscience (San Diego, CA, USA). ELISA kits for murine IL-1β and human IL-1β were purchased from Boster Biotech Co. Ltd. (Wuhan, China). MPO activity assay kit and iNOS Assay Kit were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). Immunohistochemistry kit was purchased from KeyGEN Biotech Inc. (Nanjing, China).
+ Open protocol
+ Expand
3

Isolation and Identification of Murine Immune Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols
Cells were stained in the dark on ice for 15 min with flow cytometry antibodies. Cells were then washed once with 1× PBS and resuspended in 1× PBS for sorting as described previously11 (link)–13 (link),73 (link). Antibodies used in this study were: PE anti-mouse CD45R/B220 (BD Biosciences, #553089, 1:100), PE anti-mouse TER-119 (Biolegend, #116207, 1:100), PE anti-O4 (R&D Systems, #FAB1326P, 1:100), PE anti-CD105 (eBioscience, #12–1051-82, 1:100), PE anti-CD140a (eBioscience, #12–1401-81, 1:100), PE anti-Ly-6G (Biolegend, #127608, 1:100), PerCP anti-Ly-6C (Biolegend, #128028, 1:100), APC anti-CD45 (eBioscience, #17–0451-83, 1:100), APC-Cy7 anti-CD11c (BD Biosciences, #561241, 1:100), and FITC anti-CD11b (eBioscience, #11–0112-85, 1:100). All cells were gated on the following parameters: CD105negCD140anegO4negTer119negLy-6GnegCD45Rneg. Astrocytes were subsequently gated on: CD11bnegCD45negLy-6CnegCD11cneg. Microglia were subsequently gated on: CD11bhighCD45lowLy-6Clow. Pro-inflammatory monocytes were subsequently gated on: CD11bhighCD45highLy-6Chigh. Compensation was performed on single-stained samples of cells and an unstained control. Cells were sorted on a FACS Aria IIu (BD Biosciences). For sorting of TdTomato+ astrocytes, cells were sorted according to TdTomato fluorescence judged against a wild-type control animal using a yellow-green laser on a FACS Aria IIu.
+ Open protocol
+ Expand
4

Macrophage-Mediated Phagocytosis Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols
Idelalisib was added to plated macrophages in AIM-V media and incubated at 37°C for 1 h, followed by the addition of rituximab or obinutuzumab. WIL2-S target cells were labeled with CellTracker Red (CTR; Molecular Probes; Thermo Fisher Scientific), as per the manufacturer’s protocol, and combined with the macrophages at an E:T ratio of 3:1. The cocultures were incubated for 2 h at 37°C. Cells were then stained with FITC anti-CD14 (BD Biosciences) and FITC anti-CD11b (eBioscience) and analyzed on an LSR II cytometer (BD Biosciences).
Cells within the live cell gate containing CD14+ M1 or M2c macrophages and CTR+ target cells were further separated within a dot plot quadrant. FITC+ effector macrophages were placed in the upper left quadrant of the y-axis, and CTR+ target cells were placed in the lower right quadrant of the x-axis. Double-positive cells (FITC+CTR+) in the upper right quadrant represent phagocytized target cells, and the percentage of phagocytosis was calculated as (% double-positive cells + [% double-positive cells + % target cells alone]) × 100.
+ Open protocol
+ Expand
5

Characterization of hMSCs by Flow Cytometry

Check if the same lab product or an alternative is used in the 5 most similar protocols
hMSCs were identified via flow cytometry with FITC-conjugated antibodies against FITC-anti CD90 (BD Bioscience, Franklin Lakes, NJ, USA, 1:100), FITC-anti CD45 (eBioscience, San Diego, CA, USA, 1:100), FITC-anti CD34 (eBioscience, 1:100), FITC-anti CD11b (eBioscience, 1:100), FITC-anti CD105 (eBioscience, 1:100), FITC-anti CD73 (eBioscience, 1:100), FITC-anti CD14 (eBioscience, 1:100), and FITC-anti mouse IgG as negative control.
+ Open protocol
+ Expand
6

Macrophage Immunophenotyping by Flow Cytometry

Check if the same lab product or an alternative is used in the 5 most similar protocols
APC-anti-F4/80 (17-4801-82; eBioscience), FITC-anti-CD11b (11-0112-82; eBioscience), PerCP-Cyanine5.5-anti-Ly-6C (45-5932-80; eBioscience) were used to stain marcophages ioslated from tissues. Then, FACS Aria II Cell Sorter (BD Biosciences) and FlowJo software (Tree Star) were used for FACS assay.
+ Open protocol
+ Expand
7

Quantifying Murine Macrophage Subsets

Check if the same lab product or an alternative is used in the 5 most similar protocols
The proportion of macrophage in the spleen of mice or BMDMs were analyzed by flow cytometry. These cells were stained with APC-CY7-anti-CD45, Fitc-anti-CD11b, PE-anti-F4/80, PE-CY7-anti-CD86, APC-anti-CD206, and their isotype controls (eBioscience, CA) according to the protocol. Flow cytometry was performed with the FACSCalibur (BD Immunocytometry Systems). All data were analyzed using FlowJo software.
+ Open protocol
+ Expand
8

Isolation and Purification of DAG

Check if the same lab product or an alternative is used in the 5 most similar protocols
DAG was previously isolated from S.cuneata by our laboratory  (Li et al., 2019 (link)). HPLC showed that the purity of the separated DAG was >97%. APAP was purchased from (Aladdin, Shanghai, China). APC anti-CD45, FITC anti-CD11b and PE anti-Ly6G were purchased from Invitrogen (Elabscience, Wuhan, China) for flow cytometry experiments. For detecting the concentration of liver tissue protein, BCA protein assay kit was purchased from (Thermo Scientific, Waltham, MA, USA).
+ Open protocol
+ Expand
9

Immunophenotyping of Mouse and Human Liver

Check if the same lab product or an alternative is used in the 5 most similar protocols
Livers of mice were harvested and pooled from 6-8 week C57BL/6J WT mice. Livers of human were collected from HCC patients in Shanghai Zhongshan Hospital (Shanghai, China). Red blood cells lysis was performed using ACK buffer. The panel of antibodies used in the experiments included APC-anti-F4/80 (Invitrogen, 17-4801-82), FITC-anti-CD11b (Invitrogen, 11-0112-82), APC-anti-CD11B (ThermoFisher, 11-0118-42), APC-anti-CD14 (ThermoFisher, 17-0149-42). Flow cytometry was performed using BD LSR II and BD FACS Aria III flow cytometers (BD Bioscience), and data were analyzed with FlowJo software (TreeStar, Ashland, OR).
+ Open protocol
+ Expand
10

Quantifying Colon Inflammation Biomarkers

Check if the same lab product or an alternative is used in the 5 most similar protocols
The protein levels of CD11b and S100A9 in colon tissues were detected by immunofluorescence staining as described previously [21 (link)]. In brief, colon was removed, fixed with 4% paraformaldehyde for 12 h, and cryoprotected in 30% sucrose for at least 24 h at 4 °C until the tissue pieces sank to the bottom. The colon with mucosal injury was sliced to 20 µm thick using a low-temperature slicer. The colon sections were infiltrated with 0.5% Triton X-100 in PBS, blocked with 3% BSA, incubated overnight at 4 °C with FITC-anti-CD11b (1:100, #CD11B01, Thermo Fisher) or FITC-anti-S100A9 (1:100, #73425S, Cell Signaling Technology) antibodies. The sections were counterstained with a secondary antibody (GB22302, Solarbio) for 1 h. The immunofluorescent cells were visualized and digital images were captured using a confocal laser scanning microscope.
+ Open protocol
+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required

Sign up now

Revolutionizing how scientists
search and build protocols!