Tyramide signal amplification (TSA) Plus Fluorescence Kits (PerkinElmer, Foster City, CA, USA) combined with IHC (TSA-IHC) were used for multiplex immunofluorescence staining.20 (link) Briefly, different primary antibodies were sequentially applied, followed by an incubation with horseradish peroxidase-conjugated secondary antibody and tyramine signal amplification. Nuclei were stained with 4′-6′-diamidino-2-phenylindole (DAPI, Thermo Scientific, Waltham, Massachusetts, USA) after labelling of all the human antigens. For the colocalization analysis, images were acquired with a laser confocal microscope (Olympus, Essex, UK) and analyzed with FV10-ASW Viewer software (Olympus, Essex, UK).
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Multiplex Immunofluorescence Staining Using TSA
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TSA) Plus Fluorescence Kits
4′-6′-diamidino-2-phenylindole (DAPI,
laser confocal microscope
FV10-ASW Viewer software
Antibodies
Antibody
Antigens
Dapi
Fluorescence
Horseradish peroxidase
Human
Immunofluorescence
Laser microscope
Nuclei
Tyramine
Corresponding Organization :
Other organizations : Sun Yat-sen University Cancer Center, Sun Yat-sen University
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Variable analysis
independent variables
- Primary antibodies
- Localization and expression of human antigens
- Nuclei staining with DAPI
- Positive control: Not specified
- Negative control: Not specified
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