Anti-Neutrophil Elastase antibody [EPR7479] - BSA and Azide free

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Neutrophil Elastase antibody [EPR7479] - BSA and Azide free (AB219585)

ab131260 staining Neutrophil Elastase in Human spleen tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed and paraffin-embedded, antigen retrieval was by heat mediation in Tris/EDTA buffer pH9. Samples were incubated with primary antibody (1/5000). An undiluted HRP-conjugated mouse anti-rabbit IgG was used as the secondary antibody. Tissue counterstained with Hematoxylin. PBS was used in the negative control rather than the Primary antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab131260).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Neutrophil Elastase antibody [EPR7479] - BSA and Azide free (AB219585)

ab131260 staining Neutrophil Elastase in the HL-60 cell line by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% Paraformaldehyde permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody (1/70). ab150120(1/500) an Alexa Fluor^®594-conjugated Goat anti-rabbit IgG was used as the secondary antibody. Nuclei were counterstained with DAPI.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab131260).

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Neutrophil Elastase antibody [EPR7479] - BSA and Azide free (AB219585)

Overlay histogram showing HL-60 cells stained with ab131260 (red line) at 1/20 dilution. The cells were fixed with 80% methanol. The secondary antibody used was a FITC conjugated goat anti-rabbit IgG at 1/150 dilution. Isotype control antibody (black line) was rabbit monoclonal IgG used under the same conditions. Cells also incubated without primary antibody and secondary antibody (blue line)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab131260).

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Neutrophil Elastase antibody [EPR7479] - BSA and Azide free (AB219585)

Overlay histogram showing HL60 cells stained with unpurified ab131260 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab131260, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was goat anti-rabbit Alexa Fluor^® 488 (IgG H+L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1μg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab131260).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Neutrophil Elastase antibody [EPR7479] - BSA and Azide free (AB219585)

ab131260 staining Neutrophil Elastase in Human bone marrow tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed and paraffin-embedded, antigen retrieval was by heat mediation in Tris/EDTA buffer pH9. Samples were incubated with primary antibody (1/5000). An undiluted HRP-conjugated mouse anti-rabbit IgG was used as the secondary antibody. Tissue counterstained with Hematoxylin. PBS was used in the negative control rather than the Primary antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab131260).

• WB

Lab

Western blot - Anti-Neutrophil Elastase antibody [EPR7479] - BSA and Azide free (AB219585)

This data was developed using ab131260, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

ab310335 is not suitable for detecting human cell lines in western blot assay. ab131260 and ab314916 are alternatives for detecting human cell lines.

Neutrophil elastase undergoes proteolytic cleavage to generate fragments (PMID : 25857284).

Lanes 1 - 3:

Western blot - Anti-Neutrophil Elastase antibody [EPR28386-66] (<a href='/en-us/products/primary-antibodies/neutrophil-elastase-antibody-epr28386-66-ab310335'>ab310335</a>) at 1/1000 dilution

Lanes 4 - 6:

Western blot - Anti-Neutrophil Elastase antibody [EPR7479] (<a href='/en-us/products/primary-antibodies/neutrophil-elastase-antibody-epr7479-ab131260'>ab131260</a>) at 1/1000 dilution

Lanes 7 - 9:

Western blot - Anti-Neutrophil Elastase antibody [RM1077] (<a href='/en-us/products/primary-antibodies/neutrophil-elastase-antibody-rm1077-ab314916'>ab314916</a>) at 1/1000 dilution

Lanes 1, 4 and 7:

HL-60 (human Acute Promyelocytic Leukemia promyeloblast) whole cell lysate at 20 µg

Lanes 2, 5 and 8:

THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg

Lanes 3, 6 and 9:

U937 (human histiocytic lymphoma monocyte) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 30 kDa,26 kDa

false

Exposure time: 40s

• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-Neutrophil Elastase antibody [EPR7479] - BSA and Azide free (AB219585)

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.