Biotin-Z-MAB – Mouse IgG2b with aglyco-Fc

Z-MABs have been excellent control antibodies for the assessment of the ”background bindings” of the antigen-specific, isotype matched, antibodies.  Binding of the Z-MAB antibody can be detected by the secondary antibody conjugated with enzyme, such as peroxidase or alkaline phosphatase, or fluo molecules such as FITC, Cy3, Cy5, ..etc.  With the available biotin-Z-MABs, the background binding can be determined by direct binding of the streptavidin-conjugated HRP without engaging secondary antibodies. The simplified process effectively eliminates the potentially added background signals associated with the secondary antibodies including their potential interactions with cross-species Fc receptors.  In addition, the highly stable ABC (avidin-biotin complexes) allows stringent washing conditions and leads to excellent signal to noise ratio.

A CDR-silenced mouse IgG2b antibody with an aglycosylated Fc domain (N297A), PZMU006 exhibits attenuated bindings to mouse Fcγ receptors, but its binding to FcRn is not affected.

Interacting protein(s): Fc Receptors 
Related products: Other Z-MAB^®s

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An antibody does not bind to antigen – the best control antibody commercially available.

For the control arm of an antibody study, AB Biosciences has engineered a unique antibody that does not bind to an antigen, yet it retains all other structural elements mediating all effector functions. Such a reagent represents a novel approach to unambiguously demonstrate that a specific biological activity is resulting from its binding to the cognate antigen rather than from its interactions with other proteins such as Fc receptors, complement and other Fc interacting molecules. We call this Z-MAB® for zero-binding monoclonal antibody. As a novel next generation control antibody, Z-MAB® is the only control antibody commercially available carried silenced CDR regions, that is, it is unable to bind to the antigen.

What are these control antibodies and for what applications?

Z-MAB® is a genetically engineered antibody variant whose antigen-binding capacity has been eliminated. Z-MAB® is suitable to be used as an unbiased control agent for testing specific activities of an antibody of interest.

How these control antibodies are engineered?

Starting with an antibody from a well-established antibody drug, we identified the critical amino acid residues in all 6 complementary determining regions (CDRs), engineered selected combinatorial variants and tested for the reduction/loss of their antigen-binding activities. Final candidate Z-MAB®s exhibit excellent CMC and pharmacokinetic properties.

What unmet needs these control antibodies meant to serve?

Choosing a control antibody is probably the most unscientific component for testing the specific activity of an antibody of interest. Control antibodies commercially available thus far include antibodies with no known antigens, e.g. MOPC21; antibodies raised against antigens of evolutionarily distant species, e.g. anti-KLH; antibodies reactive with a known antigen that is distinct from the antigen of interest. If cross-reactivity or high background is observed, most researchers simply move to the next control antibody available until an “ideal” control antibody is found, that is, until the expected result is observed. Z-MAB® has no active antigen-binding activity, and yet retains the overall antibody structure and conformation. It is therefore a bona fide control agent for testing antibodies.

How can these control antibodies help antibody-based research and drug development?

Our Z-MAB® product line includes the mostly commonly used isotypes of mouse, human and rat origins. For mouse Z-MAB®s, the IgG2a isotype (PZMU001) was engineered from the OKT3 (a mouse anti-human CD3 antibody) by CDR-silencing. The IgG1 isotype Z-MAB® was generated by joining the variable regions of PZMU001 to the constant regions of a mouse IgG1 antibody. Additional mouse antibody isotypes were generated using the same recombinant method. Human and rat Z-MAB®s were similarly engineered from Avastin (a humanized anti-VEGF antibody) and 53.6.72 (a rat anti-mouse CD8-alpha antibody), respectively. As such, all isotype Z-MAB®s of the same species share the same CDR-silenced variable regions. With the Z-MAB®s control agent, one no longer has to deal with the complications associated with poorly characterized “control” antibodies. In this regard, Z-MAB®s are particularly useful in animal disease models for antibody drug development where the use of unqualified control antibodies frequently leads to unexpected efficacy and for FACS analysis where the background staining frequently makes proper gating a challenge.

What control antibodies are available?

We offer a total of 16 Z-MAB®s of mouse, human and rat origin (please see the listing above). In addition to the isotype matched Z-MAB®s, Z-MAB®s carrying aglycosylated Fc domain are available as excellent control agents for antibodies whose Fc regions are not glycosylated, enzymatically or recombinantly.