Anti mouse cd45 apc

Manufactured by Thermo Fisher Scientific

Sourced in United States

The Anti-mouse CD45-APC is a monoclonal antibody that targets the CD45 antigen expressed on the surface of mouse leukocytes. It is conjugated to the APC (allophycocyanin) fluorescent dye, which can be detected using flow cytometry. This product is designed for the identification and enumeration of mouse immune cells.

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Lab products found in correlation

C tube, by Miltenyi Biotec (1 mentions) Gentlemacs dissociator, by Miltenyi Biotec (1 mentions) Anti mouse cd11b pe, by Thermo Fisher Scientific (1 mentions) Anti mouse cd11b alexa fluor 488, by Thermo Fisher Scientific (1 mentions) Facscalibur, by BD (1 mentions) Macs technique, by Miltenyi Biotec (1 mentions) Facscanto 2 flow cytometer, by BD (1 mentions) Cellsearch ctc kit, by Menarini Silicon Biosystems (1 mentions) Trypsin, by Biowest (1 mentions) Paraformaldehyde (pfa), by Ted Pella (1 mentions) Cd25 pe, by Beckman Coulter (1 mentions) Gapdh g 9, by Santa Cruz Biotechnology (1 mentions) α tubulin, by Merck Group (1 mentions) Anti human pd 1 apc, by Thermo Fisher Scientific (1 mentions) Fk506, by Merck Group (1 mentions) Dnase 1, by Roche (1 mentions) Dispase 2, by Roche (1 mentions) Collagenase type 1, by Roche (1 mentions) Cell sorter, by BD (1 mentions) Cd45 apc, by Thermo Fisher Scientific (1 mentions)

6 protocols using anti mouse cd45 apc

Flow Cytometry Analysis of Brain Immune Cells

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At 72 hours after cerebral I/R, mice were anesthetized and received transcranial perfusion with HBSS. Brain tissues from the peri‐infarct area were collected. A single‐cell suspension was prepared as described before.21, 22 For flow cytometry analysis, cells were stained on ice for 20 minutes with antimouse TLR4 Alexa Fluor 488 (1:50; eBioscience Cat#53‐9041 RRID:AB_469944; eBioscience, Inc., San Diego, CA), antimouse CD11b PE (1:150; eBioscience Cat#12‐0112 RRID:AB_465546), antimouse CD45 APC (1:150; eBioscience Cat#17‐0451 RRID:AB_469393), antimouse TLR4/MD2 complex PE (1:40; eBioscience Cat#12‐9924 RRID:AB_466264), antimouse CD11b Alexa Fluor 488 (1:100; eBioscience Cat#53‐0112 RRID:AB_469900), or antimouse CD45 APC (1:150; eBioscience Cat#17‐0451 RRID:AB_469393) and then washed with HBSS. Data were collected and analyzed on a flow cytometer (BD FACSCalibur; BD Biosciences, San Jose, CA).

Xue J., Yu Y., Zhang X., Zhang C., Zhao Y., Liu B., Zhang L., Wang L., Chen R., Gao X., Jiao P., Song G., Jiang X, & Qin S. (2019). Sphingomyelin Synthase 2 Inhibition Ameliorates Cerebral Ischemic Reperfusion Injury Through Reducing the Recruitment of Toll‐Like Receptor 4 to Lipid Rafts. Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease, 8(22), e012885.

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Isolation of Renal Collecting Duct Cells

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Viable renal intercalated cells (IC) and principal cell (PC) were enriched from IC and PC reporter mice as we previously described¹⁴. This report along with the prior observed function of collecting duct as long as 8 hour following euthanasia of the animal supports the stable properties of collecting duct cells^{14 ,22 (link),44 (link)}. Briefly, kidneys were harvested in cold PBS and immediately transferred to Accumax solution (cat.no. AM-105, Innovative Cell Technologies, CA) placed in C tubes (Miltenyi Biotec, CA). Cells were then rapidly disrupted to single cell suspension using gentleMACS dissociator (Miltenyi Biotec, CA) within 2 minutes. Renal resident immune cells separated by incubation with anti-mouse CD45-APC (Cat.no.17-0451, clone 30-F-11, eBiosciences, CA) for 30 min at 4 °C. Doublets, clumps and dead cells were removed from the cell suspension and viable cells were obtained as previously described¹⁴. Enriched ICs and PCs came from three biological replicates/sorts, each from a distinct mouse

Saxena V., Fitch J., Ketz J., White P., Wetzel A., Chanley M.A., Spencer J.D., Becknell B., Pierce K.R., Arregui S.W., Nelson R.D., Schwartz G.J., Velazquez V., Walker L.A., Chen X., Yan P., Hains D.S, & Schwaderer A.L. (2019). Whole Transcriptome Analysis of Renal Intercalated Cells Predicts Lipopolysaccharide Mediated Inhibition of Retinoid X Receptor alpha Function. Scientific Reports, 9, 545.

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Neutrophil Isolation and Priming

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Neutrophils were isolated from murine bone marrow according to Stassen et al. [12 (link)] and subsequently purified using MACS technique (Miltenyi Biotec). Purity was checked by BD FACSCanto II flow cytometer (BD Biosciences, USA) using anti-mouse CD45 APC, Clone: 30-F11 and anti-mouse Ly-6G (Gr-1) Alexa Fluor 700, Clone: RB6-8C5 antibodies (eBioscience). Neutrophils were primed according to Dewas et al. [13 (link)] by the mixture of GM-CSF and TNF-alpha (12 ng and 2.5 ng/ml respectively) for 20 min. The priming solution was enriched with 2 micromolar solution of soluble beta glucan (laminarin) as previously described [5 (link)]. Experiments were performed in complement containing medium (FCS was not heat inactivated).

Caisová V., Vieru A., Kumžáková Z., Glaserová S., Husníková H., Vácová N., Krejčová G., Paďouková L., Jochmanová I., Wolf K.I., Chmelař J., Kopecký J, & Ženka J. (2016). Innate immunity based cancer immunotherapy: B16-F10 murine melanoma model. BMC Cancer, 16, 940.

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CTC Isolation and Identification Protocol

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For human samples, 7.5 mL of whole blood was processed on the CellSearch® Autoprep system using the CellSearch® CTC kit (Menarini Silicon Biosystems), analyzed on the CellSearch® Analyzer, and assessed for the presence of CTCs as previously described [11 (link)]. For mouse samples, 50 µL of whole blood was incubated with components of the CellSearch® CTC kit including anti-EpCAM ferrofluid (25 µL), Capture Enhancement Reagent (25 µL), Nucleic Acid Dye (50 µL), Staining Reagent (50 µL), Permeabilization Reagent (100 µL), as well as anti-mouse CD45-APC (1.5 µL) (eBiosciences, San Diego, CA) as described previously [11 (link)]. Samples were manually immuno-magnetically separated and transferred to a CellSearch® MagNest™ cartridge for analysis using the CellSearch® Analyzer. In all cases, cells displaying the phenotype of EpCAM⁺/CK⁺/DAPI⁺/CD45⁻ cells with a round/oval morphology were classified as CTCs.

Kitz J., Goodale D., Postenka C., Lowes L.E, & Allan A.L. (2021). EMT-independent detection of circulating tumor cells in human blood samples and pre-clinical mouse models of metastasis. Clinical & Experimental Metastasis, 38(1), 97-108.

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Immune Cell Stimulation and Analysis

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For pharmacological stimulation, we used phorbol 12-myristate 13-acetate (PMA; Sigma) and ionomycin (Calbiochem). For antibody (Ab)-based stimulation, we used mouse antihuman or hamster antimouse monoclonal anti-CD3 and -CD28 Ab (555336, 555725, 553058, 553295; all from BD Pharmingen). For flow cytometry analyzes, we used LIVE/DEAD fixable red cell death staining kit (L23102, Invitrogen) and antimouse CD45-APC (L23102, eBioscience) or Percp-Cy5.5, CD8-BV610 and CD25-PCy7 (103132, 100744, and 102016, respectively; all from Biolegend), TCRβ-FITC (732247, Beckman Coulter) and PD-1-APC780 (47998582, eBioscience), antihuman PD-1-APC (17-9969-42, eBioscience) and CD25-PE (A07774, Beckman Coulter) and paraformaldehyde (TedPella). For western blot analysis, we used anti-DGKα (11 547–1-AP, ProteinTech), DGKζ (105195, Abcam), α tubulin (T5168, Sigma), GAPDH (G-9, Santa Cruz), horseradish peroxidase-conjugated antimouse and rabbit IgG (P0447, P0260, Dako), antirabbit IgG Dylight 800 (SA5-35571, Invitrogen) and mouse IgG AlexaFluor 680 (175775, AbcamLife). IKKβ inhibitor PS-1145 was from Sigma-Aldrich, MEK inhibitor PD98059 was from Calbiochem, and calcineurin (CaN) inhibitor FK506 was from Merck Millipore. Anti-PD-1 (nivolumab) humanized Ab was from BioVision. For TIL isolation, we used collagenase type I (Worthington), dispase II and DNase I (both from Roche). Trypsin was from Biowest.

Arranz-Nicolás J., Martin-Salgado M., Rodríguez-Rodríguez C., Liébana R., Moreno-Ortiz M.C., Leitner J., Steinberger P., Ávila-Flores A, & Merida I. (2020). Diacylglycerol kinase ζ limits IL-2-dependent control of PD-1 expression in tumor-infiltrating T lymphocytes. Journal for Immunotherapy of Cancer, 8(2), e001521.

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Isolation and Characterization of BCSPs

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Staining for BCSPs was performed using a previously described AlphaV+/CD105+/Tie2-/CD45-/CD90-/BP1- panel. Antibodies used included: AlphaV-PE (12-0512-83, eBioscience), Tie2-AlexaFluor 488 (334208, BioLegend), CD45-APC (17-0451-83, eBioscience), CD105-eFluor450 (48-1057-42, eBioscience), CD90-PE-Cyanine7 (25-0902-82, eBioscience), and BP1-Biotin (13-5891-85, eBioscience) conjugated with Streptavidin-PerCP-Cyanine5.5 (45-4317-82, eBioscience).
Staining for pericytes was performed as performed using the following antibodies: Anti-CD146-PE (12-1469-41, eBioecience), Anti-Mouse CD34-FITC (11-0341-81, eBioscience), and Anti-Mouse CD45-APC (17-0451-82, eBioscience).
Samples were incubated for 30 minutes at 4°C with an additional 15 minutes for the addition of Streptavidin- PerCP-Cyanine5.5 for the BCSPs. Following incubation samples were washed three times and filtered through a 45 micron mesh filter before being run on a FACSAria II (BD Biosciences) Cell Sorter at the University of Michigan Flow Cytometry Core. Viability was assessed by propridium iodide staining at time of sorting and analysis. Greater than 10,000 events were collected for each population. All analyses were carried out in FlowJo.
Cells from burn/tenotomy soft tissue or uninjured P4 mice were concurrently sorted to generate purified BCSPs and BCSPs-depleted subpopulations.

Agarwal S., Loder S., Li S., Shrestha S., Li J., Zhao B., Mishina Y., James A, & Levi B. (2016). Analysis of bone-cartilage-stromal progenitor populations in trauma induced and genetic models of heterotopic ossification. Stem cells (Dayton, Ohio), 34(6), 1692-1701.

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