Anti-CD33 antibody [EPR4423] - BSA and Azide free

• Flow Cyt

Lab

Flow Cytometry - Anti-CD33 antibody [EPR4423] - BSA and Azide free (AB214439)

Flow Cytometry analysis of Jurkat (Human T cell leukemia T lymphocyte, Left) / THP1 (Human monocytic leukemia monocyte, Right) labeling CD33 with ab134115 at 1/500 dilution (red). Goat anti rabbit IgG (Alexa Fluor^® 647, ab150079) at 1/2000 dilution was used as the secondary antibody. Rabbit monoclonal IgG (ab172730) was used as isotype control (Black). Unlabelled control : Cell without incubation with primary antibody and secondary antibody (Blue).

Negative control : Jurkat.
Gated on viable cells.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134115).

• IP

Unknown

Immunoprecipitation - Anti-CD33 antibody [EPR4423] - BSA and Azide free (AB214439)

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.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134115).

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] - BSA and Azide free (AB214439)

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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