Fully differentiated hSAEpCs on TWIs were fixed with 4% PFA for 15 minutes at 37°C. Inserts were washed twice with PBS for 5 min, then permeabilized by incubating in 0.2% TRITON‐X 100 at room temperature for 2 hours. Non‐specific blocking was performed by incubating inserts in PBS stain buffer containing 5% FBS and 1% BSA for 30 minutes at room temperature. Primary antibody staining was performed for 1 hour at room temperature, protected from light, using the following primary antibodies: ZO‐1 (Invitrogen Cat. # 61–7300), MUC5AC (Biotium Cat. # BNUB1134‐100), and beta‐tubulin IV (GeneTex Cat. # GTX11315) in a separate stain. Following primary antibody ZO‐1 and MUC5AC were stained with the following secondary antibodies AF647 donkey anti‐rabbit IgG (Biolegend, Cat. #406414) and FITC goat anti‐mouse IgG (BioLegend Cat. # 405305) Respectively. Separately, Beta‐Tubulin IV was detected using FITC goat anti‐mouse IgG (BioLegend Cat. # 405305). The TWI membranes were then gently cut of the inserts using a scalpel and mounted onto glass slides for imaging. Multiple Z‐Stacks were acquired using confocal imaging on a Leica TCS SPE DMi8 confocal microscope.
Goat anti mouse igg fitc
Manufactured by BioLegend
Goat anti-mouse-IgG-FITC is a secondary antibody labeled with the fluorescent dye FITC (fluorescein isothiocyanate). It is designed to detect and bind to mouse immunoglobulin G (IgG) antibodies in various immunoassays and other applications.
Automatically generated - may contain errors
Lab products found in correlation
Mouse monoclonal anti human cxcr4 antibody 12g5, by R&D Systems (2 mentions)
Tcs spe dmi8 confocal microscope, by Leica (1 mentions)
Ccl25, by R&D Systems (1 mentions)
Celltrace yellow cell proliferation kit, by Thermo Fisher Scientific (1 mentions)
Celltrace violet cell proliferation kit, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
4 protocols using goat anti mouse igg fitc
1
Immunofluorescence Staining of Airway Epithelial Cells
2
CXCR4 Expression in Leukemia Cells
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Leukemia cells (HL-60, NB4, THP-1 and U937) of 5 × 105 were incubated with mouse monoclonal anti-human CXCR4 antibody (12G5; R&D Systems, Minneapolis, MN) in BSA/PBS for 1 h at 4°C and then treated with goat anti-mouse-IgG-FITC (BioLegend, San Diego, CA). Resulting cells were subjected to C6 Accuri® flow cytometer (Accuri Cytometers, Ann Arbor, MI). The 1.5 × 104 cells were collected, acquired data were analyzed by CFlow Plus software.
3
CXCR4 Expression in Breast Cancer Cells
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The MCF-7, MCF-7/PR, SKBR-3, and SKBR-3/PR cells of × 105 were incubated with mouse monoclonal anti-human CXCR4 antibody (12G5; R&D Systems, Minneapolis, MN) in BSA/PBS for 1 h at 4˚C and then treated with goat anti-mouse-IgG-FITC (BioLegend, San Diego, CA). Resulting cells were subjected to C6 Accuri flow cytometer (Accuri Cytometers, Ann Arbor, MI), and acquired data were analyzed by FlowJo.
4
Monoclonal antibodies for immune cell analysis
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Monoclonal antibodies (mAbs) specific for mouse CD3 (17A2), CD4 (GK1.5), CD8 (53-6.7), CD19 (1D3), CD45RB (C303.16A), and integrin α4 (9C10) were all from BD Bioscience (San Jose, CA); mAb to mouse CD3 (17A2) was from Invitrogen (Carlsbad, CA); mAb to human/mouse E-Cadherin (24E10) was from Cell Signaling; mAbs to mouse F4/80 (BM8), CD25 (PC61.5), and Foxp3 (FJK-16s) were from eBioscience (San Diego, CA); mAb to mouse ICAM-1 (M-19) was from Santa Cruz (Santa Cruz, CA); mAb to integrin β7 (N1N3) was from GeneTex (Irvine, CA); rat mAb FIB504 against human/mouse β7 and rat mAb M290 against human/mouse ⍺E were prepared by using hybridomas (Developmental Studies Hybridoma Bank, University of Iowa); mAbs to mouse CD19 (1D3), human CCR9 (L053E8), FITC goat anti-rat IgG, and FITC goat anti-mouse IgG were from BioLegend; 4,6-diamidino-2-phenylindole (DAPI) was from Sigma (St. Louis, MO); CellTrace Violet Cell Proliferation Kit and CellTrace Yellow Cell Proliferation Kit were from Thermo Fisher Scientific (Waltham, MA). Human and mouse chemokines CCL21 and CCL25 were purchased from R&D Systems (Minneapolis, MN). Recombinant mouse extracellular maturation peptide E-cadherin (Asp157-Val709) fused with Fc tag were expressed in 293T cells and purified by protein A (Pierce) affinity chromatography.
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