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☑ Cell treatment (CT)
Western blot analysis of CD68 on different lysates with Rat anti-CD68 antibody (HA601596) at 1/2,000 dilution.
Lane 1: RAW264.7 (Mouse monocytic macrophage leukemia cells) cell lysate (10 µg/Lane)
Lane 2: RAW264.7 treated with PNGase F cell lysate (10 µg/Lane)
Lane 3: Mouse spleen tissue lysate (20 µg/Lane)
Lane 4: Rat spleen tissue lysate (20 µg/Lane)
Exposure time: 20 seconds; ECL: K1801
CD68 is a glycoprotein and can be deglycosylated by PNGase F. The molecular mass observed is consistent with the literature (PMID : 7680921)
Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: HA601596, 1/2,000 in primary antibody dilution buffer (K1803), overnight at 4 ℃
Secondary antibody: Goat anti-Rat IgG-HRP (HA1023), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature
Predicted band size: 35 kDa
Observed band size: 60-100 kDa
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Application: Immunofluorescence (IHC-Fr)
Species: Mouse
Tissue: Spleen
Sample: Frozen section
Antigen retrieval: Not required
Wash buffer: 1× PBST
Blocking: 10% normal goat serum + 0.5 % Triton X-100 + 0.3 M Glycine in PBS, 10 minutes at room temperature.
Primary antibody: HA601596, 1/200, overnight at 4°C.
Secondary antibody: Goat Anti-Rat IgG (iFluor™ 488, HA1133), 1.5 hours at room temperature.
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Application: Immunofluorescence (IHC-Fr)
Species: Rat
Tissue: Spleen
Sample: Frozen section
Antigen retrieval: Not required
Wash buffer: 1× PBST
Blocking: 10% normal goat serum + 0.5 % Triton X-100 + 0.3 M Glycine in PBS, 10 minutes at room temperature.
Primary antibody: HA601596, 1/200, overnight at 4°C.
Secondary antibody: Goat Anti-Rat IgG (iFluor™ 488, HA1133), 1.5 hours at room temperature.
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Application: Immunohistochemistry (IHC-P)
Species: Mouse
Tissue: Spleen
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: HA601596, 1/500, 1 hour at room temperature.
Secondary antibody: HA1023, 30 minutes 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: HA601596, 1/500, 1 hour at room temperature.
Secondary antibody: HA1023, 30 minutes at room temperature.
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Application: Immunohistochemistry (IHC-P)
Species: Rat
Tissue: Spleen
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: HA601596, 1/500, 1 hour at room temperature.
Secondary antibody: HA1023, 30 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: HA601596, 1/500, 1 hour at room temperature.
Secondary antibody: HA1023, 30 minutes at room temperature.
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☑ Relative expression (RE)
Application: Immunocytochemistry (IF-cell)
Species: Mouse
Sample: RAW264.7 (Mouse monocytic macrophage leukemia cells)/NIH/3T3 (Mouse fibroblasts)
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: HA601596, 1/500, overnight at 4°C.
Secondary antibody: Goat Anti-Rat IgG (iFluor™ 488, HA1133), 45 minutes at room temperature.
Counterstain: Beta tubulin (HA601187, red), 1/100, overnight at 4℃. The Nuclear counterstain was DAPI (Blue).
Negative control: NIH/3T3 (PMID: 29631164).
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Application: Immunofluorescence (IF-tissue)
Species: Mouse
Tissue: Spleen
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× PBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA601596, 1/200, overnight at 4°C.
Secondary antibody: Goat Anti-Rat IgG (iFluor™ 488, HA1133), 1.5 hours at room temperature.
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Application: Immunofluorescence (IF-tissue)
Species: Rat
Tissue: Spleen
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95°C.
Wash buffer: 1× PBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA601596, 1/200, overnight at 4°C.
Secondary antibody: Goat Anti-Rat IgG (iFluor™ 488, HA1133), 1.5 hours at room temperature.
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