Anti-Leukotriene A4 hydrolase/LTA4H antibody [EPR5713] - BSA and Azide free

• WB

Lab

Western blot - Anti-Leukotriene A4 hydrolase/LTA4H antibody [EPR5713] - BSA and Azide free (AB240064)

This data was developed using the same antibody clone in a different buffer formulation (ab133512).

Lanes 1-4 : Merged signal (red and green). Green - ab133512 observed at 69 kDa. Red - loading control ab8245 observed at 36 kDa.

ab133512 Anti-Leukotriene A4 hydrolase/LTA4H antibody [EPR5713] was shown to specifically react with Leukotriene A4 hydrolase/LTA4H in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266467 (knockout cell lysate ab258034) was used. Wild-type and Leukotriene A4 hydrolase/LTA4H knockout samples were subjected to SDS-PAGE. ab133512 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and 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-Leukotriene A4 hydrolase/LTA4H antibody [EPR5713] (<a href='/en-us/products/primary-antibodies/leukotriene-a4-hydrolase-lta4h-antibody-epr5713-ab133512'>ab133512</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

LTA4H knockout HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human LTA4H (Leukotriene A4 hydrolase) knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-lta4h-leukotriene-a4-hydrolase-knockout-hek-293t-cell-line-ab266467'>ab266467</a>)

Lane 3:

T-47D cell lysate at 20 µg

Lane 4:

A549 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 69 kDa

Observed band size: 69 kDa

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• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Leukotriene A4 hydrolase/LTA4H antibody [EPR5713] - BSA and Azide free (AB240064)

Overlay histogram showing Jurkat cells stained with ab133512 (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 (ab133512, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor^® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/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 antibody gave a positive signal in Jurkat cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions. This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab133512).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Leukotriene A4 hydrolase/LTA4H antibody [EPR5713] - BSA and Azide free (AB240064)

Immunohistochemical analysis of paraffin-embedded Human kidney tissue labelling Leukotriene A4 hydrolase/LTA4H with ab133512 at 1/250 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab133512).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• OI-RD Scanning

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OI-RD Scanning - Anti-Leukotriene A4 hydrolase/LTA4H antibody [EPR5713] - BSA and Azide free (AB240064)

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.