Recombinant Human Interleukin-6 Enhances the Immunoglobulin Secretion of a Rabbit–Rabbit Hybridoma

Abstract

The generation of stable rabbit-rabbit hybridomas is now possible by the recent development of a rabbit fusion partner. The ability to generate rabbit monoclonal antibodies (MAbs) can be advantageous because these rabbit immunoglobulins tend to exhibit higher affinity than murine MAbs. Furthermore, it has been observed that, in general, rabbits will elicit an immune response to antigens of limited immunogenicity in mice. Unfortunately, these rabbit-rabbit hybridomas secrete only 200 ng/mL to 5 microg/mL of immunoglobulin, which may limit larger scale production of rabbit antibodies. This study sought to determine if interleukin 6 (IL-6), which has been reported to have proliferative and secretory stimulating effects on some murine hybridomas, had any effect on a rabbit cell line that secretes a monoclonal IgG specific for estradiol. The results demonstrated that recombinant human IL-6 had a dose-dependent enhancing effect on the IgG secretion of the rabbit-rabbit hybridoma. The enhancing effect was consistent when the cells were continuously passed in the presence of IL-6. However, IL-6 did not affect the growth of the hybridoma. In contrast, no discernible effect was accomplished with recombinant mouse IL-6. Furthermore, no basal IL-6 activity was detected in the rabbit hybridoma extracellular medium. The IL-6 enhancement effect observed in this study may help to increase the immunoglobulin yield of rabbit hybridomas and to assist in the understanding of the mechanism(s) behind the lowered secretion level.

From rabbit antibody repertoires to rabbit monoclonal antibodies

“…Although this decay is frequently also observed for mouse homo-hybridomas, it appeared to be more drastic in the case of rabbit homo-hybridomas. 30, 130 For this reason, Zhu and Pytela 121 attempted to further improve the initial 240E-1 cell line by iterative subcloning to screen for clones with higher fusion efficiency, yielding hybridomas with higher genetic stability and more stable rabbit IgG secretion. The obtained fusion cell line 240E-W and its successors 240E-W2 and 240-W3, which are characterized by higher fusion efficiency and the absence of endogenous rabbit heavy- and light-chain secretion (US Patent 7,429,487), 41 are part of the proprietary RabMab platform of Epitomics, Inc. (now Abcam, Inc.) and were used to generate the therapeutic rabbit mAbs sevacizumab and APX005M discussed above.…”

“…120 As discussed above, limiting factors encountered during hybridoma technology, at least initially, included fusion efficiency and hybridoma stability. 130, 171 Also, unlike mouse hybridoma technology, rabbit hybridoma technology was patented, imposing intellectual property restrictions that have contributed to the incentive of developing alternative methods.…”

When Tissue Antigens and Antibodies Get Along

“…278,283,307 The advantages of rabbit over mouse MAbs include higher affinity (probably a result of high glycosylation), 149 suitability for use on mouse tissues without special procedures, increased specificity in some cases, and less need for antigen retrieval methods. 57,132,211,313,417 Nevertheless, in a study comparing 10 rabbit MAbs with 10 mouse MAbs targeting the same protein in animal tissues, rabbit MAbs were superior for some markers, but there was no significant overall improvement in immunoreactivity. 396 Protein A and Protein G. These cell-wall components of Staphylococcus aureus (A) and group G (G) streptococcal strains 46 are used in IHC for their high affinity for the Fc portion of IgGs.…”

Conditioned medium from gerbil—mouse T cell heterohybridomas improved antibody secretion

“…Mouse IL6 stimulates cell proliferation [ 1 , 17 , 26 ] and antibody production [ 15 , 19 ] by some mouse hybridoma cell lines. Recombinant human IL6 enhanced Ig secretion by rabbit–rabbit hybridoma cells [ 13 ]. In contrast, culture in D9(E6)C2B3-CM and D9(E4)-CM did not improve the proliferation of B11D2(C2) cells.…”