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Western blot analysis of CerS2 on different lysates with Rabbit anti-CerS2 antibody (HA724549) at 1/2,000 dilution.
Lane 1: HepG2 (Human liver cancer cell) cell lysate
Lane 2: Huh7 (Human liver cancer cell) cell lysate
Lane 3: SH-SY5Y(Human neuroblastoma cell) cell lysate
Lane 4: Jurkat (Human T-lymphoblastic cells) cell lysate
Lysates/proteins at 20 µg/Lane.
Exposure time: 1 minute; ECL: K1801
Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: HA724549, 1/2,000 in primary antibody dilution buffer (K1803), overnight at 4 °C
Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature
Predicted band size: 45 kDa
Observed band size: 40 kDa
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Application: Immunocytochemistry (IF-cell)
Species: Human
Sample: Hep G2 (Human hepatocellar carcinoma cell)
Fixation: 4% Paraformaldehyde, 15 minutes at room temperature.
Permeabilization: 0.1% Triton X-100, 15 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 1 hour at room temperature.
Antibody dilution buffer: 1% BSA in PBST.
Primary antibody: HA724549, 1/500, overnight at 4°C.
Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 45 minutes at room temperature.
Counterstain: Beta tubulin (HA601187, Red), 1/100, overnight at 4℃. The nuclear counterstain was DAPI (Blue).
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Application: Immunofluorescence (IHC-Fr)
Species: Mouse
Tissue: Liver
Sample: Frozen section
Antigen retrieval: Not required
Wash buffer: 1× TBST
Blocking: 10% normal goat serum + 1% Triton X-100 + 0.3 M Glycine in TBST, 30 minutes at room temperature.
Primary antibody: HA724549, 1/200, overnight at 4°C.
Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 1.5 hours at room temperature.
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Application: Immunohistochemistry (IHC-P)
Species: Mouse
Tissue: Liver
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724549, 1/2,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
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Application: Immunohistochemistry (IHC-P)
Species: Mouse
Tissue: Brain
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724549, 1/2,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
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Application: Immunohistochemistry (IHC-P)
Species: Human
Tissue: Brain
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724549, 1/2,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
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Application: Immunohistochemistry (IHC-P)
Species: Human
Tissue: Liver
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724549, 1/2,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
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☑ Relative expression (RE)
Application: Immunohistochemistry (IHC-P)
Species: Mouse
Tissue: Skeletal muscle
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724549, 1/2,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
Skeletal muscle exhibits low expression levels of CerS2 (PMID: 18165233).
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Application: Immunohistochemistry (IHC-P)
Species: Rat
Tissue: Brain
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724549, 1/2,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
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Application: Immunohistochemistry (IHC-P)
Species: Rat
Tissue: Liver
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724549, 1/2,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
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☑ Relative expression (RE)
Application: Immunohistochemistry (IHC-P)
Species: Human
Tissue: Skeletal muscle
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA724549, 1/2,000, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
Skeletal muscle exhibits low expression levels of CerS2 (PMID: 18165233).
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Immunoprecipitation (IP)
CerS2 was immunoprecipitated in 0.2 mg Hep G2 (Human hepatocellar carcinoma cell) cell lysate with HA724549 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA724549 at 1/2,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.
Lane 1: Hep G2 cell lysate (input)
Lane 2: HA724549 IP in Hep G2 cell lysate
Lane 3: Rabbit IgG instead of HA724549 in Hep G2 cell lysate
Exposure time: 20 seconds
Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary dilution: HA724549, 1/2,000 in primary antibody dilution buffer (K1803), 2 hours at room temperature
Predicted band size: 45 kDa
Observed band size: 40 kDa
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