anti-ERCC1 mouse monoclonal antibody, clone 4F9, 100 uL/ 30 uL

“Briefly, formalin-fixed, paraffin-embedded specimens were sliced into 4 µm sections and baked for 1 h at 65°C.. The specimens were exposed to 10 mM citrate buffer (pH 6.0) for 10 min. Tumor sections were incubated for 60 min with the mouse monoclonal anti-human ERCC1 antibody (clone, 4F9; #UM500008; OriGene Technologies, Inc., Rockville, MD, USA; dilution, 1:150), rabbit polyclonal anti-BRCA1 antibody (#AR345-5R; BioGenex Laboratories, Inc., San Ramon, CA, USA; dilution, 1:50), rabbit monoclonal anti-RRM1 antibody (clone, EP242; #AC-0217RUO; Epitomics, Abcam, Cambridge, UK; dilution, 1:100) and mouse monoclonal anti-TUBB3 antibody (clone SDL3D10; #MU177-UC; BioGenex Laboratories, Inc.; dilution, 1:50).. The tissue sections were incubated with polyclonal goat anti-rabbit (#ab150077; Abcam; dilution, 1:200) and goat anti-mouse (#ab150115; Abcam; dilution, 1:200) IgG biotinylated secondary antibodies for.. “

“Blocking of unspecific protein–antibody interactions was performed with 20% human AB serum in PBS for 1 h at room temperature.. Primary antibody for ERCC1 was the new highly specific monoclonal anti-mouse antibody (mAb) clone 4F9 (UM500008, dilution 1:100) that was purchased from OriGene Technologies (Rockville, MD, USA).. A mouse negative control was used (Dako North America).”

“nutes to 958C, for antigen retrieval. Slides were cooled to room temperature and incubated in a peroxidase blocking solution (Dako, Carpinteria, California) for 10 minutes. Slides were washed and incubated with the primary antibodies (ERCC1 and 4F9, OriGene, Rockville, Maryland; ERCC1 and D6G6, Cell Signaling Technology, Danvers, Massachusetts) for 1 hour at room temperature. Several iterations of antigen retrieval, primary antibody dilution, and incubation times were tested to determine the opti”.

” the cells completely. The 96 crude cell lysates were transferred to a 96-deep well plate with 2 ml capacity (Axygen) and spun at 4000 x rpm to remove insoluble cellular debris. The clarified supernatants were transferred to a 96-well v-shape plate (BD) and stored at -80C for future protein microarray printing. The recombinant protein expression for every overexpression lysate used for microarray printing was validated by Western-blot analysis with the anti-DDK antibody (OriGene, TA50011-100). “