Abcam
Anti-CD32B antibody - C-terminal (ab151497)
ab151497
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Browse Similar Products: CD32 Flow Cytometry Antibodies
Detection of the three forms of the low affinity Fc gamma receptor CD32 in human cells is complicated by the near identity of the extracellular domains. CD32B, the inhibitory FcgRIIb, has a unique intracellular domain with an ITIM. Antibodies directed against the intracellular C-terminal domain should theoretically distinguish this receptor from CD32A or CD32C. I tested three antibodies: ab151497 (Abcam, rabbit polyclonal), ab77093 (Abcam, goat polyclonal) and F-4 (Santa Cruz, mouse monoclonal). The antibodies were tested for use in flow cytometry on PBMC, using basophils as a positive control and T cells as a negative control. All antibodies provided signal in flow cytometry. All antibodies non-specifically stained T cells as well as basophils. Ab151497 provided the largest signal to noise ratio, but in an unconjugated form will not be useful for determining absence of expression.
Flow Cytometry
PBMC
1:100 for 30min at 4C in BD permeabilization buffer on fixed/permeabilized PBMC
2.5% FCS (can't block Fc receptors here)
1:200 Zenon rabbit IgG labeling reagent AlexaFluor 647 (Invitrogen/ThermoFisher)
none
BD FACSCanto LSR flow cytometer
Surface staining with FITC-conjugated pan-CD32 antibody FLI8.26 revealed no staining for CD32 on CD3+ T cells, but bright staining on CCR3+FceRI+ basophils. Intracellular staining with ab151497 anti-CD32B showed bright staining on CD3+ T cells and brighter staining on CCR3+FceRI+ basophils. Ab151497 also brightly stained all mouse leukocytes, including those from FcgRIIb-deficient mice. The other CD32B antibodies tested showed similar profiles, with less difference between CD32B-expressing and CD32B-non-expressing cell types. Specificity of staining with ab151497 may be improved by blocking the cells extensively with non-specific rabbit IgG and then using directly fluorophore-conjugated ab151497.
None
Detects CD32B, does work in flow cytometry
Majority of signal is non-specific.
Not very useful in an unconjugated format, but may be useful if directly conjugated.