Hybridoma fusion cell lines contain an aberrant kappa transcript☆
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Cited by (111)
Humanised recombinant antibody fragments bind human pancreatic islet cells
2018, Journal of Immunological MethodsCitation Excerpt :For the VL chains, PCR products were obtained with two Vκ primers for each, and sequencing revealed that only one in each pair was productive. The identical, non-productive sequences showed premature termination due to a frameshift mutation at the Vκ-Jκ junction and has been associated with the hybridoma fusion partner's parent myeloma line (Carroll et al., 1988). Each productive VH and VL sequence was used to search the international ImMunoGeneTics database (IMGT/LIGM-DB, http://imgt.cines.fr) (Giudicelli et al., 2006) in order to identify the mouse VH and VL genes utilized, and to correct the 5′ sequence modified by the forward primer.
Identification and verification of hybridoma-derived monoclonal antibody variable region sequences using recombinant DNA technology and mass spectrometry
2017, Molecular ImmunologyCitation Excerpt :Although universal degenerate primers simplify and accelerate the molecular cloning process, their use can often lead to the amplification of contaminating and aberrant sequences via incorporation of the erroneous nucleotides at degenerate sites in the primer (Bialon et al., 2014). Moreover, different myeloma fusion partners inherently transcribe different aberrant and contaminating sequences making a single solution difficult to achieve (Cochet et al., 1999; Carroll et al., 1988; Kohler and Milstein, 1975; Kearney et al., 1979). Our in silico analysis of IgG1 proteins identified highly conserved sequence in Ig leader, constant and hinge regions that allowed us to define optimal PCR primers and reaction conditions to limit off-target binding and successfully amplify Ig VR sequences from all hybridoma cell lines tested (Tables 1 and 2).
A novel monoclonal antibody suitable for the detection of leukotriene B4
2017, Biochemical and Biophysical Research CommunicationsCitation Excerpt :Sequencing data confirmed these findings: VL Sequences deduced from experiments with primer sets for kappa light chains illustrated exclusively non-productive aberrant kappa transcripts from myeloma cells. One of them was already published to occur in hybridomas after fusion with sp2/0-Ag14 myeloma cells by Carroll et al. [23]. A continuous VL transcript sequence in contrast could only be generated with lambda specific primer sets (Table 2).
Anti-tumor activity of human IgG1 anti-gp75 TA99 mAb against B16F10 melanoma in human FcgammaRI transgenic mice
2014, Immunology LettersCitation Excerpt :mRNA levels of aberrant light chain can be high and may even exceed levels of the light chain mRNA of TA99. Most aberrant light chain transcripts are not translated into functional light chains, because they contain pseudo genes or reading-frame shifts [23,29]. However, some aberrant light chain mRNAs can be translated to protein and secreted when combined with heavy chains [30].
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This work was supported by USPHS grant CA 33399 to Dr Ronald Levy. W.L. Carroll is a recipient of an NIH clinical investigator award CA 01059. Computer resources used to carry out this study were provided by the National Institutes of Health-sponsored BIONET National Computer Research for Molecular Biology (grant number RR-01685-05).