Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free

8 product images

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  Western blot - Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free (ab240001)

  This data was developed using the same antibody clone in a different buffer formulation (Anti-SP1 antibody [EPR6662(B)] ab124804).
  

  Lanes 1-2: Merged signal (red and green). Green - Anti-SP1 antibody [EPR6662(B)] ab124804 observed at 100 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 37 kDa.

  Anti-SP1 antibody [EPR6662(B)] ab124804 Anti-SP1 antibody [EPR6662(B)] was shown to specifically react with SP1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line Human SP1 knockout HeLa cell line ab265519 (knockout cell lysate Human SP1 knockout HeLa cell lysate ab257698) was used. Wild-type and SP1 knockout samples were subjected to SDS-PAGE. Anti-SP1 antibody [EPR6662(B)] ab124804 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4°C at 1 in 5000 and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

  All lanes: Western blot - Anti-SP1 antibody [EPR6662(B)] (Anti-SP1 antibody [EPR6662(B)] ab124804) at 1/5000 dilution

  Lane 1: Wild-type HeLa cell lysate at 20 µg

  Lane 2: SP1 knockout HeLa cell lysate at 20 µg

  Lane 2: Western blot - Human SP1 knockout HeLa cell line (Human SP1 knockout HeLa cell line ab265519)

  Performed under reducing conditions.

  Predicted band size: 81 kDa

  Observed band size: 100 kDa

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  Flow Cytometry (Intracellular) - Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free (ab240001)

  Intracellular Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling SP1 with Purified Anti-SP1 antibody [EPR6662(B)] ab124804 at 1/100 dilution (10 μg/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue). This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-SP1 antibody [EPR6662(B)] ab124804)

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  Immunocytochemistry/ Immunofluorescence - Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free (ab240001)

  Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling SP1 with Purified Anti-SP1 antibody [EPR6662(B)] ab124804 at 1:200 dilution (4 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with None. Goat anti rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) was used as the secondary antibody at 1:1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
  This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-SP1 antibody [EPR6662(B)] ab124804)

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free (ab240001)

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human gastric carcinoma tissue sections labeling SP1 with Purified Anti-SP1 antibody [EPR6662(B)] ab124804 at 1:2000 dilution (0.41 µg/ml). Perform heat mediated antigen retrieval using Antigen Retrieval Buffer (100X Tris-EDTA Buffer, pH 9.0) ab93684 (Tris/EDTA buffer, pH 9.0). Purified)ImmunoHistoProbe one step HRP Polymer (ready to use) was used for detection. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
  This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-SP1 antibody [EPR6662(B)] ab124804)

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free (ab240001)

  Anti-SP1 antibody [EPR6662(B)] ab124804, at 1/100 dilution staining SP1 in paraffin-embedded Human colon tissue, by Immunohistochemistry.
  

  This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-SP1 antibody [EPR6662(B)] ab124804).

  Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

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  OI-RD Scanning - Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free (ab240001)

  We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody.
  Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.

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  ChIC/CUT&RUN sequencing - Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free (ab240001)

  ChIC/CUT&RUN was performed using a pAG-MNAse at a final concentration of 700 ng/mL. 2.5X10^5 of Human wild-type HeLa cell line (ab255928) or Human SP1 knockout HeLa cell line (Human SP1 knockout HeLa cell line ab265519) were used along with 5µg of Anti-SP1 antibody [EPR6662(B)] ab124804 [EPR6662(B)]. Assay Quality Control was conducted using 5µg Anti-CTCF (Anti-CTCF antibody [EPR18253] - ChIP Grade ab188408) on the same cell lines. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads.? The negative IgG control Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730 is also shown.
  
  Additional screenshots of mapped reads can be found in the Protocol booklet in the Support and downloads section.
  
  The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.
  
  This data was developed using the same antibody clone in a different buffer formulation (Anti-SP1 antibody [EPR6662(B)] ab124804).

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  Western blot - Anti-SP1 antibody [EPR6662(B)] - BSA and Azide free (ab240001)

  SP1 Western blot staining using rabbit Anti-SP1 antibody

  This data was developed using Anti-SP1 antibody [EPR6662(B)] ab124804, the same antibody clone in a different buffer formulation.
  

  Western blot: Rabbit Monoclonal[EPR6662(B)] to SP1 Anti-SP1 antibody [EPR6662(B)] ab124804 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 90 kDa in Wild-type MCF7 cell lysates with no signal observed at this size in SP1 knockout MCF7 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc Milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

  All lanes: Western blot - Anti-SP1 antibody [EPR6662(B)] (Anti-SP1 antibody [EPR6662(B)] ab124804) at 1/1000 dilution

  Lane 1: Wild-type MCF7 at 20 µg

  Lane 2: SP1 knockout MCF7 at 20 µg

  Lane 2: Western blot - Human SP1 knockout MCF7 cell line (ab287770) at 20 µg

  Lane 3: HeLa at 20 µg

  Lane 4: Ramos at 20 µg

  Secondary

  All lanes: Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

  Performed under reducing conditions.

  Predicted band size: 81 kDa

  Observed band size: 90 kDa