Anti-CD64 antibody [EPR4623]

• Flow Cyt

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Flow Cytometry - Anti-CD64 antibody [EPR4623] (AB109449)

ab109449 at 1/100 dilution staining CD64 in U937 cells by Flow cytometry (shown in red). Rabbit IgG negative control (shown in green).

• WB

Lab

Western blot - Anti-CD64 antibody [EPR4623] (AB109449)

False colour image of Western blot : Anti-CD64 antibody [EPR4623] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab109449 was shown to bind specifically to CD64. A band was observed at 45-50/55-80 kDa in wild-type THP-1 cell lysates with no signal observed at this size in FCGR1A knockout cell line ab275843 (knockout cell lysate ab275817). To generate this image, wild-type and FCGR1A 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 (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-CD64 antibody [EPR4623] (ab109449) at 1/1000 dilution

Lane 1:

Wild-type THP-1 cell lysate at 20 µg

Lane 2:

FCGR1A knockout THP-1 cell lysate at 20 µg

Lane 2:

Western blot - Human FCGR1A knockout THP-1 cell line (<a href='/en-us/products/cell-lines/human-fcgr1a-knockout-thp-1-cell-line-ab275843'>ab275843</a>)

Lane 3:

U937 cell lysate at 20 µg

Lane 4:

HEK-293 cell lysate at 20 µg

Predicted band size: 43 kDa

Observed band size: 45-50 kDa

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• WB

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Western blot - Anti-CD64 antibody [EPR4623] (AB109449)

All lanes:

Western blot - Anti-CD64 antibody [EPR4623] (ab109449) at 1/1000 dilution

Lane 1:

U937 cell lysate at 10 µg

Lane 2:

THP1 cell lysate at 10 µg

Lane 3:

HL60 cell lysate at 10 µg

Predicted band size: 43 kDa

Observed band size: 64-78 kDa

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• OI-RD Scanning

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OI-RD Scanning - Anti-CD64 antibody [EPR4623] (AB109449)

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.