Animal experiments were performed in accordance with a protocol approved by the Institutional Animal Care and Use Committee at MDACC. Molm14 cells (0.6 × 106) stably expressing a dual luciferase-GFP reporter (Molm14-GFP/Luc) were injected via the tail vein into NOD/SCID IL2Rγ Null-3/GM/SF (NSGS) mice (Jackson Laboratory, Bar Harbor, ME). Once engraftment was confirmed by the IVIS-200 noninvasive bioluminescence in vivo imaging system (Xenogen, Hopkinton, MA), mice were either untreated or treated with VS-4718 twice a day at 75 mg/kg via oral gavage (n = 10/group) for 16 days. Leukemia burden was monitored by IVIS in vivo imaging, flow cytometric measurement of human CD45 cells (anti-human CD45 antibody, BD Biosciences) in mouse PB, and immunohistochemical staining for human CD45+ cells in mouse tissues (stained with anti-human CD45 antibody and visualized by Biotin-free Tyramide Signal Amplification System, both from Dako, Carpinteria, CA). Mouse survival was recorded.
Anti human cd45 antibody
Manufactured by Agilent Technologies
Sourced in United States
The anti-human CD45 antibody is a laboratory reagent used to detect and identify cells expressing the CD45 surface antigen. CD45 is a transmembrane protein tyrosine phosphatase that is expressed on the surface of most hematopoietic cells, including lymphocytes, monocytes, and granulocytes. The antibody can be used in various cell analysis techniques, such as flow cytometry, to distinguish and characterize different cell populations.
Automatically generated - may contain errors
Lab products found in correlation
Anti human cd45 antibody, by BD (1 mentions)
Biotinylated horse anti mouse igg, by Vector Laboratories (1 mentions)
Discovery xt processor, by Roche (1 mentions)
Dab detection kit, by Roche (1 mentions)
Permount, by Thermo Fisher Scientific (1 mentions)
Ab64238, by Abcam (1 mentions)
Celld imaging 3, by Olympus (1 mentions)
Autostainer link 48, by Agilent Technologies (1 mentions)
Bx51 microscope, by Olympus (1 mentions)
4 protocols using anti human cd45 antibody
1
Efficacy of VS-4718 in Xenograft AML Model
2
Immunohistochemical Analysis of CD45 in Mice
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The fixed harvested tissues that were obtained from the sacrificed mice were embedded in paraffin and 7-µm slices were cut using a microtome (RM 2155; Leica, Germany). Slices were then immunohistochemically stained with anti-human CD45 antibody (#M0701; Dako, Carpinteria, CA, USA) and incubated for 60 min with biotinylated horse anti-mouse IgG (#MKB-2225B; Vector Laboratories, Newark, CA, USA). Immunohistochemical staining was performed at the Molecular Cytology Core Facility at Memorial Sloan Kettering Cancer Center in New York, NY, USA, using a Discovery XT processor (Ventana Medical Systems, Oro Valley, AZ, USA). Detection was performed using a DAB detection kit (Ventana Medical Systems) according to manufacturer instructions. Slides were counterstained with hematoxylin and coverslipped with Permount (Thermo Fisher Scientific).
All instruments and equipment used in the study were calibrated to ensure accurate measurements and reliable results.
All instruments and equipment used in the study were calibrated to ensure accurate measurements and reliable results.
3
Immunohistochemical Analysis of CD45
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Spleen and sternum from recipients were collected, incubated at least 24 h in 3.7% neutral buffered formalin (pH 7.4), embedded in paraffin and sectioned at 8 μm onto Superforst-Plus slides. Samples were incubated with an anti-human CD45 antibody (Dako, #M0701) in Dako Autostainer Plus solution, then incubated with a sheep anti-mouse IgG conjugated to HRP and processed as per standard protocols using 3,3’ Diaminobenzidine staining (#ab64238, Abcam) for all immunoblots, as previously described101 . Samples were imaged and annotated using an Aperio ScanScope and software system (Leica Biosystems).
4
Histochemical Analysis of Organ Infiltration
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Sections of paraffin-embedded samples of infiltrated (liver, spleen, hindlimbs and backbone) and normal (lung, heart and kidney) organs were hematoxylin and eosin (H&E) stained and the presence of toxicity was analyzed. Moreover, in order to detect AML cells in infiltrated tissues, immunohistochemical analysis with anti-human CD45 antibody (DAKO) was done in paraffin-embedded tissue samples. Staining was performed in a Dako Autostainer Link 48, following the manufacturer's instructions. Two independent observers evaluated all samples, using an Olympus BX51 microscope (Olympus). Images were acquired using an Olympus DP72 digital camera and processed with CellD Imaging 3.3 software (Olympus).
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