Xp. Li et al., HYBRIDOMA SCREENING USING AN AMPLIFIED FLUORESCENCE MICROASSAY TO QUANTIFY IMMUNOGLOBULIN CONCENTRATION, Hybridoma, 14(1), 1995, pp. 75-78
The early identification of antibody-secreting hybridomas significantl
y reduces the time and resources devoted to unproductive colonies, To
quantify the immunoglobulin concentration in hybridoma supernatants wi
thin 2 weeks of fusion, we used immunomagnetic microspheres to capture
immunoglobulin in the hybridoma culture supernatant, The captured imm
unoglobulin was detected using a goat anti-mouse second antibody liked
to beta-galactosidase. With data transformation to correct for the no
nlinear accumulation of the fluorescent reaction product, the enzymati
c hydrolysis of fluorescein digalactoside permitted the reliable detec
tion of less than 10 pg of immunoglobulin per milliliter, To determine
the value of quantifying immunoglobulin concentration within 2 weeks
of fusion, the amplified fluorescence microassay was applied to the ev
aluation of 3 consecutive fusions and more than 1200 growing hybridoma
colonies, Using antibody concentrations greater than 10 ng/ml as a th
reshold for routine subculture, the selection of hybridoma colonies ba
sed on antibody secretion was threefold more efficient than selection
based on colony growth alone, These results suggest the utility of the
early determination of immunoglobulin concentration in the selection
of hybridoma colonies.