All tags Monoclonal RabMAb® technology: the need, the attempts and the breakthrough

The RabMAb story

RabMAb® technology: the need, the attempts and the breakthrough

​​

The need

​​​The rabbit immune system is complex and generates highly sensitive antibodies. Due to their superior antigen recognition, antibodies that are difficult to make in mice can often be produced in rabbits (1, 2). 

The need to generate rabbit monoclonal antibodies was addressed 20 years ago, as polyclonals have the drawback of inconsistent signal quality (i.e. non-specificity, high background). In contrast, monoclonals offer unique epitope recognition, lot-to-lot consistency and sustainable supply.

 “With increasing demands from scientific communities and healthcare industries for antibodies with better sensitivity and specificity, rabbit monoclonals were the answer.

Weimin Zhu, Co-inventor of RabMAb® technology, Abcam – 

It’s long been known that rabbit antibodies are highly sensitive and could be generated against targets that aren’t easily achievable in mouse (1, 2). The reason for this is that rabbit is a more complex animal compared to mouse and therefore has a more complex immune system. However, until around 20 years ago, only polyclonal antibodies in rabbit were achievable. As polyclonals have the drawback of inconsistent signal quality (i.e. non-specificity, high background), there was a need for developing monoclonals in rabbit. A monoclonal version offers the benefits of unique epitope recognition, lot-to-lot consistency and sustainable supply.
 “With increasing demands from scientific communities and healthcare industries for antibodies with better sensitivity and specificity, rabbit monoclonals were the answer.”
Weimin Zhu – 

The attempts

Many attempts were made to generate rabbit monoclonal antibodies following the development of mouse hybridoma technology, where myeloma cells were fused with mouse spleen cells (3).

The first of these were mouse-rabbit heterohybridomas, which were not ideal as they were difficult to clone, unstable and had a somewhat limited production source. (4). 

In 1995, Katherine Knight and her colleagues at Loyola University of Chicago developed a double transgenic rabbit that over-expressed the oncogenes v-abl and c-myc. These were under the control of the immunoglobulin heavy and light chain enhancers, and the rabbit formed a myeloma-like tumor.

This enabled the the isolation of a plasmacytoma cell line, named 240E-1, the first fusion partner to generate rabbit monoclonals. Fusion of 240E-1 cells with rabbit lymphocytes produced hybridomas that secreted rabbit monoclonal antibodies in a consistent manner, but the stability of the fusion partner was a concern (5).

The breakthrough

​At the University of California San Francisco in 1996, Weimin Zhu and Robert Pytela obtained 240E-1 from Dr. Knight’s laboratory and developed the technology further (6). 

240E-1 cells were repeatedly subcloned and selected to improve the characteristics of the fusion line. Selection criteria included fusion efficiency, growth, and morphological characteristics (i.e. bright appearance under a phase contrast microscope).

Selected subclones were further analysed to ensure they produced stable hybridomas and effeciently secreted monoclonal antibodies. After multiple rounds of subcloning, a new cell line named 240E-W was generated that showed greater fusion efficiency and stability.

This was followed by the development of RabMAb technology, Abcam's patented technology to develop highly specific monoclonal antibodies with high sensitivity.

Cell line 240E-W has since been further modified to eliminate endogenous IgG issues. Since 2006, the new fusion partner (240E-W2, a core part of the RabMAb technology as we know it today), has been used for production of rabbit monoclonal antibodies for both research and commercial applications. 

Today

RabMAb technology makes it possible to develop high quality monoclonal antibodies for research, diagnostic and therapeutic applications.In the past 16 years, RabMAb technology has proven to be the premium technology to fulfill various needs within the scientific community.

Alongside our industry leading catalog of over 6,000 RabMAb products, Abcam is pleased to offer its powerful RabMAb technology for the creation of sensitive, specific antibodies on a custom basis.

References

  • 1. Krause RM. Experimental approaches to homogenous antibody populations: factors controlling the occurrence of antibodies with uniform properties. Fed Proc. 1970;29:59-65. 
  • 2. Weller A, Meek J, Adamson ED. Preparation and properties of monoclonal and polyclonal antibodies to mouse epidermal growth factor (EGF) receptors: evidence for cryptic EGF receptors in embryonal carcinoma cells. Development. 1987;100:351-363.
  • 3. Raybould TJ, Takahashi M. Production of stable rabbit-mouse hybridomas that secrete rabbit mAb of defined specificity. Science. 1988;240:1788-1790.
  • 4. Verbanac KM, Gross UM, Rebellato LM, et al. Production of stable rabbit-mouse heterohybridomas: characterization of a rabbit monoclonal antibody recognizing a 180 kDa human lymphocyte membrane antigen. Hybridoma. 1993;12:285-295.
  • 5. Spieker-Polet H, Sethupathi P, Yam PC, et al. Rabbit monoclonal antibodies: generating a fusion partner to produce rabbit-rabbit hybridomas. Proc Natl Acad Sci U S A. 1995;92:9348-9352.
  • 6. Liguori MJ, Hoff-Velk JA, Ostrow DH. Recombinant human interleukin-6 enhances the immunoglobulin secretion of a rabbit-rabbit hybridoma. Hybridoma. 2001 Jun;20(3):189-98.