FLAG-tagged PTP1B was detected with anti-FLAG antibody (20543-1-AP [Proteintech]). PTP1B was detected with anti-PTP1B clone FG6-1G (MABS197; Merck Millipore) or with clone H-135 (sc-14021) from Santa Cruz Biotechnology to detect specifically the C-terminal part. Anti-CD22 antibody was purchased from Novus Biological (clone 2H1C4), anti-CD79A (Igα; clone HM47) and anti-SYK (4D10.2) from BioLegend. The antibodies directed against; BTK-pY223, #5082; PLCγ1, #5690; PLCγ2, #3872; PLCγ2-pY759, #3874; SYK-p525/526, #2710; ERK, #4695; ERK-pT202/Y204, #4370; GRB2, #3972; and GAPDH, #2118 were all from Cell Signaling. Anti-phospho-CD22-pY807 (ab32040) antibody was purchased from Abcam. Anti-Myc-tag (66004-1-Ig), anti-APLP2 (15041-1-AP) and anti-CBL (25818-1-AP) were from Proteintech. Anti-BTK (sc-1107) and anti-BCAP (AF4857) antibodies were obtained from Santa Cruz Biotechnology and R&D Systems Inc., respectively. All antibodies were used according to the manufacturer’s instructions. For imaging, a ChemoCam (Intas) equipped with a full-frame 3.2 megapixel Kodak KAF-3200ME camera was used. Western blot signals were quantified with Quantity One 4.6.9 (Bio-Rad). For statistical analysis, two-sample t tests were performed over three replicates using Microsoft Excel 2013. Error bars represent the SEM.
Anti flag antibody 20543 1 ap
Manufactured by Proteintech
Sourced in China
The Anti-FLAG Antibody (20543-1-AP) is a protein detection tool used to identify and locate FLAG-tagged proteins in various applications. It is a highly specific antibody that recognizes the FLAG epitope, which is a short amino acid sequence commonly used as a protein tag. The antibody can be utilized in techniques such as Western blotting, immunoprecipitation, and immunohistochemistry to detect and study FLAG-tagged proteins.
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Lab products found in correlation
Pt202 y204 erk, by Cell Signaling Technology (1 mentions)
Quantity one 4, by Bio-Rad (1 mentions)
Plcγ2, by Cell Signaling Technology (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
Plcγ1, by Cell Signaling Technology (1 mentions)
Syk p525 526, by Cell Signaling Technology (1 mentions)
Ripa lysis buffer, by Beyotime (1 mentions)
Protease inhibitor, by Roche (1 mentions)
Enhanced bca protein assay kit, by Beyotime (1 mentions)
Anti ha antibody h6908, by Merck Group (1 mentions)
Tritc conjugated phalloidin, by Merck Group (1 mentions)
Ki 67 antibody, by Cell Signaling Technology (1 mentions)
E cadherin, by BD (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Sangon (1 mentions)
N cadherin, by Cell Signaling Technology (1 mentions)
3 protocols using anti flag antibody 20543 1 ap
1
Western Blot Antibody Detection Protocol
2
Protein Extraction and Western Blot Analysis
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Proteins were extracted with RIPA lysis buffer (P0013B, Beyotime, Shanghai, China) containing protease inhibitor (4693159001, Roche, Indianapolis, USA) and quantified by an enhanced BCA protein assay kit (P0009, Beyotime, Shanghai, China). Western blot analysis was performed according to a previously described standard protocol [23] (link). Anti-FOXM1 antibody (20459) was purchased from Cell Signaling Technology (Boston, USA). Anti-hTERT antibody (32020) was purchased from Abcam (Cambridge, UK). Anti-tubulin antibody (AF1216) was purchased from Beyotime. Anti-HA antibody (H6908) was purchased from Sigma-Aldrich (St. Louis, USA). Anti-Flag antibody (20543-1-AP) and anti-GST antibody (3G12B10) were purchased from Proteintech (Wuhan, China). These primary antibodies were used at a 1:1,000 dilution for immunoblotting analysis.
3
Immunofluorescence and Immunohistochemical Staining Protocol
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Cells were seeded on 1% gelatin coated coverslips in 6-well plates, and then fixed in 4% paraformaldehyde for 15 minutes at room temperature (RT), permeabilized in 0.2% Triton X-100 for 3 minutes at RT, blocked in blocking buffer (PBST with 5% BSA) for 1 hour at RT. Following removing blocking buffer, cells were incubated with anti-cofilin antibody (BS2183; Bioworld Technology), or anti-Flag antibody (20543-1-AP; Proteintech Group) or E-cadherin (562869, BD Biosciences, San Jose, CA), or N-cadherin (13116, Cell Signaling technology) in blocking buffer for over-night at 4℃. Secondary antibodies were Alexa Fluor 488-conjugated mouse IgG (115-545-166; Jackson ImmunoResearch Laboratories, West Grove, PA) and Alexa Fluor 488-conjugated anti-rabbit IgG (111-545-144; Jackson ImmunoResearch Laboratories). For phalloidin staining, cells were incubated with TRITC-conjugated phalloidin (p1951; Sigma-Aldrich, St. Louis, MO) and DAPI (1 mg/mL working concentration) (D6584; Sangon Biotech, Shanghai, China) for 2 hours at RT. For Ki67 immunohistochemical staining, tumor tissues were embedded in paraffin, sectioned in 6-μm thickness, and stained with ki67 antibody (9129; Cell Signaling Technology) using UltraSensitiveTM SP (Rabbit) IHC Kit (KIT-9707; Fuzhou Maixin Biotechnology, Fuzhou, China).
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