Orthotopic allografts were generated as in the efficacy studies. When tumors reached 200–400 mm3 (about 2 weeks post-cell injection), R848-Toco/HA-Toco (25 μg on R848 basis) in 50 μL volume was injected intratumorally. After 24 h, tumors were dissected and embedded in OCT medium (Fisher Scientific), followed by storage at −80 °C. Tumors were sectioned (10 μm) with a cryotome. Sections were fixed for 2 × 10 min with acetone and washed with PBS. Primary antibodies were diluted to 5 μg/mL in blocking buffer (5% goat serum in PBS) and incubated overnight at 4 °C. Antibodies used were Alexa Fluor® 488 anti-CD8a, Alexa Fluor® 594 anti-CD11b, and Alexa Fluor® 647 anti-CD11c (BioLegend). After antibody staining, sections were stained with DAPI (0.5 μg/mL in PBS; Invitrogen) for 10 minutes, mounted in SouthernBiotech™ Fluoromount-G™ Slide Mounting Medium and stored in the dark at 4 °C. Images were acquired using an Olympus IX-81 inverted epifluorescence microscope at 10x magnification. and images were captured with a Hamamatsu EM-CDD Digital Camera. The entire tumor section was imaged. Many 10x images were montaged together using SlideBook 6 to view the whole section.
Alexa fluor 647 anti cd11c
Manufactured by BioLegend
Alexa Fluor® 647 anti-CD11c is a fluorescently labeled antibody that binds to the CD11c cell surface marker. CD11c is expressed on the surface of dendritic cells, macrophages, and other antigen-presenting cells. This antibody can be used to identify and study these cell populations in flow cytometry and other applications.
Automatically generated - may contain errors
Lab products found in correlation
Alexa fluor 488 anti cd8a, by BioLegend (2 mentions)
Alexa fluor 594 anti cd11b, by BioLegend (2 mentions)
Fluoromount g slide mounting medium, by Southern Biotech (1 mentions)
Oct medium, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Oxaliplatin, by Merck Group (1 mentions)
Succinic anhydride, by Merck Group (1 mentions)
α tocopherol, by Merck Group (1 mentions)
N n diisopropylethylamine dipea, by Merck Group (1 mentions)
Tetrabutylammonium hydroxide solution tbaoh, by Merck Group (1 mentions)
1 bis dimethylamino methylene 1h 1 2 3 triazolo 4 5 b pyridinium 3 oxide hexafluorophosphate hatu, by Chem-Impex (1 mentions)
3 3 dimethylaminopropyl 1 ethyl carbodiimide hydrochloride edc hcl, by Chem-Impex (1 mentions)
Vt1200, by Leica (1 mentions)
Alexafluor647 anti cd3, by BioLegend (1 mentions)
Anti cx3cr1 pe, by BioLegend (1 mentions)
Triton x 100, by Merck Group (1 mentions)
Fv3000, by Olympus (1 mentions)
3 protocols using alexa fluor 647 anti cd11c
1
Intratumoral Injection and Immunofluorescence
2
Synthesis and Characterization of Cisplatin-Hyaluronate Conjugates
3
Immunofluorescent Staining of Mouse Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Fresh mouse tissues were soaked in 4% paraformaldehyde (PFA) for 10 hours and then sectioned at a thickness of 100 μm by a vibrating blade microtome (Leica VT1200, Germany). Then, slices were blocked in PBS containing 0.02% Triton X-100 (Sigma, Missouri, USA) and 1% bovine serum albumin (BSA) for 1 hour and then incubated with a directly-labeled antibody overnight at 4 °C. Primary antibodies used in this study were as follows: PE anti-CX3CR1 (Clone SA011F11 Cat# 149005, BioLegend), Alexa Fluor 647 anti-CD11c (Clone N418 Cat# 117312, BioLegend), and Alexa Fluor 647 anti-CD3 (Clone 17A2 Cat# 100209, BioLegend). For indirectly-labeled antibody staining, the slices were blocked as described above, then incubated with goat anti-mouse VAChT (Clone ABN100, Sigma) for 1 hour at room temperature, and further stained with the secondary antibody donkey anti-goat IgG(H + L) (Clone A21447, Biolegend) overnight at 4 °C. After washing off the unbound antibody, samples were enclosed using PBS containing 50% glycerol (w/w). Samples were imaged under an inverted confocal fluorescence microscope (FV3000, Olympus, Japan) with dry 10× NA0.4 and dry 20× NA0.8 objectives.
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