Antibody Preparation

mAbs are prepared by:
Monoclonal Antibody Core Laboratory
Interdisciplinary Center for Biotechnology Research
University of Florida
2033 Mowry Road
Gainesville, FL 32610

mAbs are prepared using the following components:
CELLine 350 bioreactor flask from Integra Biosciences
BD Cell MAb basal medium from Becton Dickinson

Super low IgG fetal bovine serum from HyClone

mAb lots are prepared as follows:
Each lot of mAb is prepared from one bioreactor flask.
Each flask is started with a vial of hybridoma cells stored at -80°C.
Typically, 10-12 harvests of 8-10 mL each are obtained from one flask over 10-12 weeks.
Harvests are sterile filtered, adjusted to 0.02% sodium azide, and stored at 4°C.
Octet quantitation of IgG yield is done on every other harvest.

IgG concentrations are typically in the range of 0.75 to 1.5 mg/mL.
Harvests are pooled, sterile filtered, and stored at 4°C (pool = lot).

mAbs are tested as follows:
mAbs are tested for recognition of antigen of the correct molecular weight by western blotting.
Western blot strips are prepared from yeast proteins separated by SDS-PAGE.
Blots are probed with mAbs diluted 1/1000 in PBS + 0.1% Tween 20.
Results are visualized by chemiluminescence.

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