Optimize, Adapt and Overcome - 23

Optimize, Adapt, and Overcome: Tips from the Cell Line Development Front * Enhanced Development of Virus-Specific Hybridomas Using ClonePix and CloneSelect Imager Technologies

CloneSelect™ Imager technology. This highly
efficient automated clone selection aided rescue
and stabilization of a high-titer hybridoma cell
line producing a highly specific antibody to an
immunogenic viral antigen. The pairing of
technologies provided sufficient material for
immunodiagnostic device development.

Materials
* ClonePix System
* CloneSelect Imager
* CloneMedia Semi-Solid Media for
hybridomas/myelomas
* CloneDetect Reagent, Mouse IgG (H+L)
Specific, Fluorescein.

Rescuing Poorly Producing Hybridomas
Traditional methods for hybridoma development
involve chemically-mediated fusion of a myeloma
cell line with the splenocytes of a host animal
immunized with an antigen of interest. The fused

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hybridomas undergo notoriously slow and
inefficient selection processes by limiting dilution,
followed by ELISA screening to isolate a limited
number of antigen-specific monoclonal hybridoma lines. With an extended timeline (up to 30
weeks) and increased handling, limiting dilution
methods produce lower titer cell lines where
the vitality and productivity of the cell line could
never be precisely elucidated.
The first ds-DNA viral specific hybridoma cell line
was developed in 1990s using limiting dilution
methods, thus the monoclonality and stability
was not optimized. Repeated cultures (static and
roller) were done over the years with varying sera
and media compositions, yet the clones were
rendered unstable (Figure 1). Moreover, antibody
yields never reached more than 3 mg/L range and
the quality of the purified IgG was never uniform
with varying amounts of aggregation observed
in the preps.
The goal was to rescue and stabilize the specific

hybridoma cell line by sub-cloning a higher titer
antibody producing parental line. ClonePix
Platform offers a significant technological
advantage utilizing a semi-solid methylcellulose
media and label-free detection by CloneDetect
Reagent to re-screen a very large number of
previously fused hybridomas (Figure 2).
High throughput screening methodology using
a ClonePix System enabled the rescue of stable,
higher producing, stable clones from the lower
producing parental clones.

Rapid Isolation of High-Yielding
Target Clones Secreting Antibody
ClonePix Systems have demonstrated significant
improvements in workflow productivity and
cost-effectiveness, increasing the probability of
finding rare secretors, thereby reducing the time
required for monoclonal antibody generation
by up to 50% as compared to limiting dilution
methods.

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Optimize, Adapt and Overcome

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