PMC:4419340 / 37758-38747 JSONTXT

{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/4419340","sourcedb":"PMC","sourceid":"4419340","source_url":"https://www.ncbi.nlm.nih.gov/pmc/4419340","text":"The TIRF assay.\nThe TIRF assay was performed on the transfected HEK-293T cells as described in “Cell purification and culture.” V5 tag–IL-17RC WT and mutant (Q138*, R376*, and R378*) vectors were constructed at the N terminus by the pcDNA3.1 Directional TOPO Expression kit (Life Technologies). After being fixed in 3.7% paraformaldehyde, the cells coated glass coverslips and were incubated with anti-V5 antibody (Life Technologies) and Phalloidin (Life Technologies) for 3 h at room temperature. After being washed, the glass coverslips were incubated with anti–mouse Alexa Fluor 488 (Life Technologies) for 1 h at room temperature and then stained with DAPI (Sigma-Aldrich). Fluorescence data were acquired with the Eclipse Ti-E TIRF imaging system (Nikon). Images were recorded with the QuanTEM 512 SC camera (Roper Scientific) and acquired with NIS-Elements AR software (version 3.1; Nikon). Image sets were processed with ImageJ version 1.43 software (National Institutes of Health).","divisions":[{"label":"Title","span":{"begin":0,"end":15}}],"tracks":[]}