Stages of the Bacterial Growth Curve
Knowing the difference between the lag, log, stationary, and death phases of a cell growth curve may sound like biology basics but applying that knowledge in active cell culture is a sign of a more fundamental understanding of microbiological processes. The subtle nuances of cell culture that the uninitiated may overlook can help those of us paying close attention to optimize growth conditions or successfully scale up production with less headache and heartache. While it’s important to know what they are, it’s more important to know their secrets.
Lag Phase
This initial cell growth phase is generally characterized by metabolic activity but not growth. The cells utilize media to synthesize the small molecules necessary for replication. This stage is where the cells start to increase in size but not necessarily in number. Stopping a culture during the lag phase may seem counterintuitive but if the primary interest is learning about the first metabolic activity cells experience, this may in fact be the prime time to finish. Understanding the physiological and regulatory pathways responsible for reproduction is an important element that may be learned from cellular activity in the lag phase. Despite the potential wealth of information available from this stage, it is the least understood and researched of them all.
Log (Exponential Growth) Phase
Once cells enter the exponential growth or log phase, it’s all about the numbers. This is when cells divide by binary fission and the doubling of each generation creates the exponential growth for which the phase is named. Metabolic activity during this stage is high but generally limited to the necessary steps required for reproduction. The log phase may be extended by additional supplementation of nutrients equal to the increasing growth rate. This is known as an exponential fed-batch cell culture and requires constant monitoring of cell growth conditions as well as advanced knowledge of when and how much to feed. If the number of cells produced is the final product then stopping cell culture at the end of the log phase is important. This is especially true in industrial microbiology where cost and time affect the bottom line and may mean the difference between profits or losses.
Stationary Phase
As environmental conditions change and become the limiting factor in growth, cells slow reproduction and enter a stationary phase. In this stage cells are technically still reproducing but it is less than before and roughly equal to the amount of cell death. The stationary phase is when conditions are no longer ideal for reproduction and cells start to experience stress that results in changed metabolic activity. Depending on the type of cell being grown, this may be the stage where protein production and excretion take place or where spore-forming bacteria start to produce endospores allowing them to survive harsh conditions. During the stationary phase of cell culture, it is generally considered to be the products of the cells that are of interest, and that makes this an important part of bioprocessing. Additional supplements may be added to keep the cells alive but stressed enough to continue producing rather than reproducing. In cases where cell culture may last days, weeks, or months, like the production of monoclonal antibodies via hybridoma cells, or the production of ethanol and CO2 via yeast fermentation, the stationary phase is where the magic happens.
Death Phase
As waste builds up and nutrient-rich media is depleted, the death phase is the point where the living cells stop metabolic functions and begin the process of death. As cells lyse and fill the culture with what was once on their insides, the environment changes one last time and exponential decay begins. While it’s said that all good things must come to an end, the death phase of cell culture can oftentimes be the most important stage of bioprocessing. This is true in cases where cells lyse and release amino acids, proteins, polysaccharides, and free fatty acids that may be the intended products. Food sciences are especially affected by the death phase in cell culture and will often control the exact time it begins by altering environmental conditions to stop the metabolic cellular activity. In bioprocessing and cell culture, it’s understood by all those involved that the little things can make a big difference, and in a field so filled with questions, having even some of the answers can be the pathway to success rather than failure. Knowing what’s happening to your cells at each stage of their life cycle may just give you that edge you need.