Producing recombinant antibodies works in various prokaryotic and eukaryotic systems. Each system yields its own pros and cons. In this quick guide to expression of recombinant antibodies, we look at the mammalian expression and outline the various antibody formats.
Expression Host Cell Line
Recombinant antibody expression can use a variety of host cell lines, including yeast, insect, bacterial (E. coli), plant, and mammalian. Mammalian is one of the most commonly used host cells because it offers proper expressing and folding and native-like post-translational modification yields. But it’s worth noting that each host cell will result in variable outcomes.
Two mammalian hosts that researchers often use include the Chinese Hamster Ovary (CHO) and Human Embryonic Kidney (HEK239) because they offer stable and transient expressions and work relatively well for recombinant antibody expressions.
Designing the Expression Vector
Researchers will conduct an initial comprehensive analysis of the primary sequence, indicating the level of challenge surrounding the target production. This, and the various available bioinformatic tools, help design a viable expression vector.
After designing the target and expression vectors, creating a cloning scheme is essential. It’s common for researchers to develop a high throughput expression and cloning process using a parallelization strategy such as Ligation Independent Cloning or In-Fusion Cloning, also known as recombinant-based cloning.
Transient vs. Stable Expressions
Transient and stable expressions require the target gene to integrate with the host cell. But one method may be better than the other based on your yield requirements.
Transient transfection is an excellent option for those who need results quickly or with a minimum yield requirement. During transient transfection, the target DNA cannot integrate with the host cell and may lose itself through cell division.
Compared to transient transfections, a stably transfected cell can and will become one with the host genome, which results in adequate replications. Stable expressions begin as a transiently expressed cell, but through amplification, a transient transfection becomes stable and results in stable clones. Stable expression is suitable for large-scale production measures and yields a longer setup time.
Purification Methods
The chosen purification method for recombinant antibody expression can improve overall production. There are two methods available: manual purification or a chromatography system. The following purification methods outline which applications they may serve best:
Affinity chromatography: High recovery rates, high selectivity, and high resolution.
Size exclusion chromatography: Allows for increased purity and sample homogeneity.
Ion exchange chromatography: Separates antibodies based on surface charges.
Hydrophobic interaction chromatography: Viable in intermediate or polishing requirements during purification.
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