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REAfinity™ Recombinant Antibodies  - Flow cytometry is in their genes

  • 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 hybridoma derived, 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.

 

 

Made to meet new reproducibility standards

Get reliable results through consistent antibody quality

The manufacturing process of REAfinity™ Antibodies ensures high performance consistency based on the following aspects:

  1. REAfinity Antibodies are derived from a defined DNA sequence, which encodes only one type of heavy and light chain, ensuring high antibody purity (fig. 2).
  2. Production under biologically and chemically defined, standardized culture conditions in mammalian cells, resulting in high lot-to-lot consistency (fig. 3).
  3. Analytical, biochemical, and cell-based testing of every manufactured lot (fig. 3).


Figure 1: Mass spectrometry analysis of purified antibodies shows that REAfinity Antibodies are defined products, while hybridomagenerated antibodies can be a mixture. (A) Mass spectrometry analysis of two examples of hybridoma-generated monoclonal antibodies shows that both contain a second light chain with a molecular weight of approx. 23,635 Da. In one example bottom left, the amount of the contaminating light chain was by far exceeding the productive light chain of approx. 24,142 Da. (B) Two examples of recombinant REAfinity Antibodies show pure light chain populations.

Figure 2: Staining performance of REAfinity Antibodies is nearly identical between different lots. Two manufacturing lots of five different CD56 REAfinity Antibody conjugates were compared by flow cytometry using the MACSQuant Analyzer. The two histogram curves (black and red) represent the two lots. Lot-to-lot staining performance was nearly identical, when using human peripheral blood mononuclear cells (Human PBMCs) from a single donor.

Hassle-free analysis – no more background signal

Analyze what matters

All REAfinity™ Antibodies contain a specifically mutated human IgG1 Fc region that abolishes their binding to Fcγ receptors (fig. 4). This allows for background-free analysis and eliminates the need for additional blocking steps such as using a FcR blocking reagent (fig. 5 and 6)

Figure 3: Staining with REAfinity Antibodies shows no background signal, even without FcR block. Human PBMCs were stained with either a mouse monoclonal CD158a-PE antibody or REAfinity CD158a-PE (clone:REA284). Staining was performed without (top) and with (bottom) FcR block. The mouse monoclonal antibody binds unspecifically to CD56-negative cells (A). Staining with the REAfinity Antibody, shows no background signal Cells were analyzed by flow cytometry on the MACSQuant® Analyzer 10. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide (PI) fluorescence.

Straightforward experiment planning with only one isotype control

Save efforts when setting up experiments

All REAfinity™ Antibody Clones contain the same specifically mutated human IgG1 sequence as their constant region. This offers the possibility to work with only one type of isotype control, reducing the complexity of experiment planning and saving time. Hybridoma-derived monoclonal antibodies on the other
hand, are composed of antibody isotypes derived from different species and therefore require multiple isotype controls. This complicates not only the management of reagent inventory, but also panel design when setting up experiments.


Figure 4: REAfinity Antibodies reduce the complexity of panel design with only one isotype control required.

Optimized for flow cytometry applications

The perfect reagents for your cell analysis

REAfinity™ Antibodies are designed with flow cytometry in mind. The following properties ensure their superior performance as compared to traditional hybridoma-derived monoclonal antibodies:

1. Greater purity
2. No background signal
3. Highly optimized and standardized fluorochrome conjugation process


Figure 5: Specific detection of human CD116+ cells with clone REA 211. Human PBMCs were stained with either a PE-conjugated mouse monoclonal antibody (A) or PE-conjugated REAfinity Antibody (B) recognizing CD116. Cells were also stained with CD14-FITC. Cells were analyzed by flow cytometry on the MACSQuant® Analyzer 10. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide (PI) fluorescence.

Flexibility in flow cytometry with thousands of recombinant antibodies



REAfinity Recombinant Antibody clones are available for hundreds of specificities. Search our list for your antibody of interest:



All REAfinity clones



They are conjugated to a variety of fluorochromes to address multicolor flow cytometry needs. Current options include traditional dyes, like FITC, PE, APC, and Miltenyi Biotec´s proprietary Vio® Dyes:



Figure 1: Emission spectra of VioDyes: VioBlue, VioBright 515, Vio515, VioGreen, VioBright FITC, PE-Vio615, PerCP-Vio700, PE-Vio770, APC-Vio770

Videos

REAfinity™ Recombinant Antibodies – behind the scences
3min

Reproducibility is a cornerstone of science. Watch the behind-the-scenes story of how we developed REAfinity Recombinant Antibodies to address today's reproducibility crisis and pave the way for new reproducibility standards.
 
Webinar: Recombinant antibodies for improved flow cytometry
49min

Join Dr. Kalpana Singh and her webinar on the advantages of recombinantly generated REAfinity antibodies that can enhance the reproducibility of your results and improve flow cytometry analysis
 
Documentary: Antibody validation challenges

38min

The documentary highlights the concerns raised by key stakeholders to understand if this recent awareness in reproducibility is truly a crisis or an opportunity to lay down some guidelines that will strengthen our faith in scientific data and outcomes going forward.
 

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