Anti-CD33 antibody [EPR4423] - Low endotoxin, Azide free

• IP

Unknown

Immunoprecipitation - Anti-CD33 antibody [EPR4423] - Low endotoxin, Azide free (AB215383)

This IP data was generated using the same anti-CD33 antibody clone, EPR4423, in a different buffer formulation (cat# ab134115).

ab134115 (purified) at 1/60 immunoprecipitating CD33 in 10 μg THP-1 (Lanes 1 and 2, observed at 67-75 kDa). Lane 3 - PBS. For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution. Blocking buffer and concentration : 5% NFDM/TBST Dilution buffer and concentration : 5% NFDM/TBST

All lanes:

Immunoprecipitation - Anti-CD33 antibody [EPR4423] (<a href='/en-us/products/primary-antibodies/cd33-antibody-epr4423-ab134115'>ab134115</a>)

Predicted band size: 40 kDa

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• WB

Supplier Data

Western blot - Anti-CD33 antibody [EPR4423] - Low endotoxin, Azide free (AB215383)

False colour image of Western blot : Anti-CD33 antibody [EPR4423] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab134115 was shown to bind specifically to CD33. A band was observed at 60-80 kDa in wild-type THP-1 cell lysates with no signal observed at this size in CD33 knockout cell line ab273831 (knockout cell lysate ab273785). To generate this image, wild-type and CD33 knockout THP-1 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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 IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution. This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134115).

All lanes:

Western blot - Anti-CD33 antibody [EPR4423] (<a href='/en-us/products/primary-antibodies/cd33-antibody-epr4423-ab134115'>ab134115</a>) at 1/1000 dilution

Lane 1:

Wild-type THP-1 cell lysate at 10 µg

Lane 2:

Western blot - Human CD33 knockout THP-1 cell lysate (ab273785) at 10 µg

Lane 3:

HL-60 cell lysate at 10 µg

Lane 4:

Jurkat cell lysate at 10 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 40 kDa

Observed band size: 60-80 kDa

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• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-CD33 antibody [EPR4423] - Low endotoxin, Azide free (AB215383)

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.