Products and Services - MACS Flow Cytometry - Reagents - Antibodies and dyes - REAfinity Antibodies

Recombinant antibodies for flow cytometry

  • High purity and lot-to-lot consistency for greater reproducibility
  • Eliminates tedious and costly Fc receptor-blocking steps
  • One universal isotype control for convenience and cost savings

REAfinity™ Antibodies are recombinant antibodies that provide superior lot-to-lot consistency and purity compared to mouse or rat monoclonal antibodies. They've been recombinantly engineered to produce highly specific antibodies that require no FcR blocking step. Additionally, they all have the same IgG1 isotype.

 

Hundreds of REAfinity specificities are available. Choose from a variety of fluorochromes to address your multicolor flow needs – FITC, VioBright™ FITC, PE, APC, VioBlue®, VioGreen™, PE-Vio770™, APC-Vio770™, PerCP-Vio700™.

Curious if a REA clone is available for your specificity/target of interest? View all REA clones here. Have questions about REAfinity Antibodies? See the frequently asked questions page for more info.

 

We back up our claims that REAfinity™ Antibodies are the best antibodies available with real data. They are demonstrated to be highly specific, result in better staining than mouse or rat monoclonals, and without the need of Fcγ receptor blocking step.

  • Highly specific: better staining than mouse and rat monoclonals
  • Clear resolution of target population: enables straightforward data analysis
  • High stain index: due to an optimized fluorochrome labeling process and low background

Figure 1: Specific detection of CD158b2+ cells with REA147. PBMCs were stained with a PE-conjugated REAfinity Antibody (A) and mouse monoclonal PE-conjugated antibody (B) recognizing CD158b2. Additionally, staining with CD56-APC (# 130-100-698) was performed followed by flow cytometry analysis on the MACSQuant® Analyzer. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide (PI) fluorescence.

Figure 2: Specific recognition of CD116+ cells with REA211.  PBMCs were stained with a PE-conjugated
REAfinity Antibody (A) and mouse monoclonal PE-conjugated antibody (B) recognizing CD116. Additionally, staining with CD14-FITC (# 130-080-701) was performed followed by flow cytometry analysis on the MACSQuant® Analyzer. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide (PI) fluorescence.

Figure 3: Specific detection of Dectin-1+ cells with REA154. Bone marrow cells from BALB/c mice were stained with a PE-conjugated REAfinity Antibody (A) and a mouse monoclonal PE-conjugated antibody (B) recognizing Dectin-1. Additionally, staining with CD11b-APC (# 130-102-493) was performed followed by flow cytometry analysis on the MACSQuant® Analyzer. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide (PI) fluorescence.

REAfinity™ Antibodies have specifically mutated human IgG1 Fc regions. These mutations virtually eliminate unspecific binding to Fcγ receptors, as demonstrated in figure 4. Staining with REAfinity Antibodies therefore makes the use of FcR blocking reagents redundant resulting in time and cost savings.
 

Figure 4: Immunoassay comparing the binding of a REAfinity™ Antibody (light orange) and a mouse monoclonal antibody (orange) recognizing CD144 to the five cellular Fcγ receptors. The binding of immobilized Fcγ receptors to REAfinity Antibodies and mouse monoclonal antibodies was detected using secondary antibodies conjugated to HRP.

REAfinity™ Antibodies show only minimal background signals

In the absence of an FcR blocking reagent, staining with the mouse monoclonal antibody results in strong non-specific background signals (figure 5A). Therefore, it is necessary to include an FcR blocking step prior to staining with the mouse monoclonal antibody (figure 5B). In contrast, staining with the REAfinity Antibody allows the specific detection of the CD158a+ target population, even without FcR blocking reagent (figure 5C).
 

Figure 5: REAfinity Antibodies show low background signals. PBMCs were stained with a PE-conjugated REAfinity Antibody (C) and a mouse monoclonal PE-conjugated antibody (A, B) recognizing CD158a. Additionally staining with CD56-FITC (# 130-100-746) was performed followed by flow cytometrical analysis on the MACSQuant Analyzer. The staining with the mouse monoclonal antibody conjugate was performed either with (A) or without (B) the pre-treatment with FcR Blocking Reagent, whereas no FcR blocking reagent was included prior to staining with the REAfinity Antibody (C). Cell debris and dead cells were excluded from the analysis based on scatter signals and PI fluorescence.

One universal isotype control

One universal isotype control

REAfinity™ Antibodies all have human IgG1 as their isotype. The result is that you need only one type of isotype control for all REA clones, named REA Control Antibodies or clone REA293. This control is available in two formats – one for surface- and one for intracellular-expressed antigens.

What does one isotype control per fluorochrome mean to you?
  • You will experience significant cost savings since you will need only one control.
  • You will save time spent on searching for the antibody isotype.

High purity - no Ig chain mixtures

Mouse monoclonal antibodies often contain mixtures of different IgG heavy and light-antibody chains (figure 6). This impurity can lead to lot-lot variations and could impact the reproducibility of your experiments. 

REAfinity™ Antibodies offer several major advantages over mouse monoclonal antibodies for long term studies, including:

  1. REAfinity Antibody products contain only contain one heavy and one light-chain for higher lot-to-lot consistency (figure 6).
  2. The unmatched quality of REAfinity Antibodies also arises from its manufacture in mammalian cells and production under a highly controlled manufacturing process.
  3. Compared to the average mouse monoclonal antibody, REAfinity Antibodies offer greater purity and higher lot-to-lot consistency to make REAfinity Antibodies your reagent of choice.

Figure 6: Mass spectometry analysis of mouse monoclonal (A) and REAfinity Antibody samples (B). A mixture of antibody chains can be observed in the case of the mouse monoclonal antibody while the REA clone only shows one heavy chain (50 kDa) and one light chain (25 kDa).

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Miltenyi Biotec Inc.
Phone: +1 800 FOR MACS
Fax:+1 877 591 1060
macs@miltenyibiotec.com
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