Antibody drugs are developed through the QbD method for upstream production processes, utilizing the advantages of experimental design (DoE) and high-throughput parallel bioreactors technology and equipment to efficiently complete antibody laboratory scale research and development processes. 

Early understanding of the relationship between key quality attributes (CAQs) and key process parameters (CPP) of the product, as well as determining the design space in the research and development process.

Once these small-scale R&D processes are completed, the next step is to scale up the processes. Ideally, efficient process scaling can complete the transition from R&D scale to pilot/production scale within 2-4 months. How to achieve efficient process transfer and scaling up is a severe test of the experience and level of the entire process technology team.

The goal of process amplification is to ensure stable growth of cells in a relatively constant environment for product expression as the cell culture scale gradually expands.

The measurement criteria include cell density, growth rate, viability, product expression rate, and glycosylation level, among others. The key control parameters in the amplification process of cell culture technology can be divided into two types.

One is volume independent, such as temperature, DO, and pH. Another type is influenced by volume and geometric dimensions, such as stirring speed and ventilation flow rate.

Due to the diverse range of tank suppliers used during the research and development phase, the material (disposable or glass), aspect ratio, stirring blade diameter, and tank diameter ratio of the tank, as well as the enlarged tank used, are generally not completely consistent; 

Even tanks of different volumes from the same supplier cannot achieve proportional size enlargement. This poses a great challenge for determining parameters such as stirring and ventilation after amplification.

To maintain consistency in the cultivation environment, developers typically adopt the following amplification criteria.

In the cell culture process, the shear force of the stirring blade is one of the important factors to consider. Different engineering cell lines have varying tolerance to shear forces. 

Early CHO cells had lower tolerance to shear forces, while current engineering cell lines have greatly improved their tolerance to shear forces.

Shear force is often reflected through blade tip velocity. 

The diameter and speed of the stirring blade determine the magnitude of the blade tip velocity. Due to the requirements of tank design, the increase in tank volume will also result in an increase in the diameter of the stirring blade. 

Therefore, in the mode of constant blade tip velocity, the stirring speed of large volume tanks is lower than that of small volume tanks. 

The amplification of constant leaf tip velocity allows cells to grow under the same shear stress environment. This amplification strategy is suitable for small-scale amplification and production.

Mixing time is a relatively simple criterion for amplification. Especially in the chemical industry, the constant mixing time can be directly used as a basis for amplification. 

However, in cell culture processes, small-scale (such as 2L or less) volumes can quickly achieve mixing time, while larger volumes require higher leaf tip velocities to achieve mixing time. This will lead to an increase in shear force, causing damage to the cells.

KLa represents the rate at which oxygen enters the liquid phase from the gas phase. The appropriate KLa in the reactor is the key to process amplification, and O2, as an important nutrient for cells, affects their normal growth and metabolism. 

The constant KLa amplification criterion provides cells with the same oxygen transfer environment.

However, the determination of KLa is influenced by many factors. For example, during the cultivation process, factors such as stirring speed and ventilation flow rate require a lot of testing and analysis to determine the appropriate KLa. 

In practical operation, KLa increases with the increase of working volume. After reaching a certain volume, KLa CO2 will interact with KLa again, increasing the difficulty of constant amplification of KLa.

P/V is related to many factors such as stirring power (Np, Power Number), tank diameter, stirring blade diameter, working volume, liquid density, etc., which to some extent reflect the degree of mixing and affect the mixing and mass transfer of the culture system. 

Therefore, constant P/V is recommended as a criterion for many process amplifications and is currently the most commonly used amplification strategy. Considering the different tolerance of cell shear forces, the common P/V range is 10-40 W/m3.

In addition to the above four amplification criteria, when scaling up to a certain volume, it is also necessary to consider the negative effects of pCO2 on cell growth and protein expression. 

When cultivating in small volumes, due to the fact that the introduced gas can carry away most of the CO2 produced by cell metabolism during the ascent process, there is basically no need to consider CO2 stripping.

In large volume cultivation, KLa CO2 decreases with the expansion of reactor scale, that is, the CO2 removal capacity decreases. The gas saturation and bubble volume in the system directly affect CO2 removal, and increasing the aeration rate and bubble residence time can accelerate removal. 

Some reactor suppliers have integrated CO2 removal functionality into the control system to facilitate the control of pCO2.

With the continuous improvement of antibody technology, the challenges faced by process amplification are also constantly changing. Many times, process developers need to flexibly adjust the amplification process based on the different characteristics of cell lines and the different expressed products. 

More often than not, multiple factors such as P/V, blade tip velocity, CO2 removal, etc. are comprehensively considered to obtain the optimal scaling up production process while ensuring CQAs.