Cmparison with PEG method in hybridoma preparation
|
|
hybridoma
growth supplement |
hybridoma
positive rate |
antibody-production
positive rate |
|
GenomONE™ - CF
EX |
- |
38/96 (40%) |
9/96
(9%) |
|
+ |
96/96
(100%) |
96/96 (100%) |
|
PEG1500 |
- |
3/96 (3%) |
1/96
(1%) |
|
+ |
36/96
(38%) |
9/96 (9%) |
GenomONE - CF
EX
produces more antibody secreting
hybridomas than PEG. (following cell fusion, cells were
grown in media containing hybridoma growth supplement as
indicated).
Normal BALB/c mouse splenocytes (1x10^8 cells) not
sensitized with antigen were fused to X63-Ag8.653
myeloma cells (1x10^7 cells) using GenomONE(tm)-CF
EX or
PEG1500. The fused cells obtained with each agent were
inoculated onto five 96-well plates (Day 0). Beginning
the following day, half of the culture medium (10%FBS/RPMI1640)
was replaced with HAT medium at five points of time (Days
1, 2, 3, 5, and 8), and the growth of colonies in each
well was assessed on Days 10 -11 to determine the
hybridoma-positive rate
(an indicator of efficiency of fusion). On Day 12, mouse
antibody level (IgG + IgA + IgM) in the supernatant was
measured by ELISA, to calculate the antibody
production-positive rate. The effect of adding a
commercially available hybridoma supplement to the
medium after fusion was also assessed (supplement was
also added to the HAT medium).