Download Corning® cellgro® hybriGRO SF™ Animal-Free

Corning® cellgro® hybriGRO SF™
Animal-Free, Serum-Free Defined Medium
With Corning® cellgro® glutaGRO™ Supplement
Adaptation in Corning cellgro hybriGRO SF medium
Maintenance in Corning cellgro hybriGRO SF medium
Either direct adaption or weaning techniques can be used to
adapt cells to hybriGRO SF medium. While most hybridoma
cells do not require weaning, this method is recommended as
a precaution due to possibly significant differences in formulation between the current media used and hybriGRO SF
medium.
Hybridoma cultures should be split into fresh, serum-free
medium every 3-4 days and seeded at 1 to 2 x 105 cells/mL
to maintain optimal growth and productivity.
Direct Adaptation
1. A seeding density of double the normal seeding density
for the cell line is suggested for the first passage.
2. Subculture cells when the cell density is 2 × 106 to 3 × 106
cells/mL.
3. For passage numbers 2-4, subculture the cells to a viable cell density of 2 × 105 cells/mL in fresh, serum-free
medium.
Weaning
Gradually change the medium over 2-3 passages by blending
the two media 50:50 for the first passage and then 25:75 for
the second. Seed cells at ~ 5 x 105 cells/mL.
Cat. No.
Unit Size
Storage
40-215-CV
500 mL
2-8°C
Thawing
1. Once the last amount of ice thaws, quickly transfer the
contents of the vial so that the cells are seeded at 4 to
5 x 105 cells/mL in fresh serum-free medium. If using
centrifuge take care to use low speed as cells are extremely
fragile following freezing.
2. Monitor cell growth and viability and resume normal
maintenance schedule.
100
3.50E+06
90
3.00E+06
80
70
2.50E+06
60
hybriGRO SF
DMEM + 10% serum
hybriGRO SF
DMEM + 10% serum
50
40
2.00E+06
1.50E+06
30
1.00E+06
20
5.00E+05
10
0
1
2
3
4
5
0.00E+00
Time (days)
Results
Comparison of hybriGRO SF medium versus standard serum-supplemented medium
using AE1 (ATCC HB-72™) hybridoma cell line. Growth is improved with hybriGRO SF
medium, with comparable viability.
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1. Cells must be in the mid-logarithmic phase of growth
with viability >90% prior to adaptation.
2. Higher seeding density than normal is recommended to
increase conditioned medium carryover early on during
adaptation.
1. Gently pellet mid-log phase cells for freezing. Remove
supernatant and resuspend in a volume of cryopreservation medium (e.g. 500 µL conditioned medium: 500 µL
fresh medium + 50-100 µL DMSO) to obtain a final cell
density of 1 to 5 x106 cells/mL.
2. Dispense into cryovials and freeze following standard
procedures.
Cell Viability (%)
Important factors leading to successful adaptation include:
Cryopreservation