Cell culture is a fundamental practice in biological research, involving the growth of cells in a controlled laboratory environment. This allows scientists to study cell behavior, physiology, and molecular processes in a simplified setting. To maintain the health and continuous growth of these lab-grown cells, a routine procedure known as “passaging cells” is performed.
Understanding Cell Passaging
Cell passaging, also referred to as subculturing, is the process of transferring cells from one culture vessel to a new one containing fresh growth medium. This procedure becomes necessary as cells proliferate and reach a certain density, often referred to as confluence, where they cover the available growth surface. At this point, the cells may experience stress due to overcrowding, depletion of nutrients in the culture medium, and accumulation of metabolic waste products.
Preventing overgrowth is important because prolonged confluence can lead to changes in cell behavior, health, and even cell death. By diluting the cell population into new vessels, fresh nutrients are supplied, waste products are removed, and adequate space for continued proliferation is provided. Passaging allows for the expansion of cell populations, ensuring a continuous supply of healthy and consistent cells for various experiments. It also helps maintain cells in an optimal growth phase for research.
The Steps of Cell Passaging
The process of passaging cells begins with observing the cell culture under a microscope to determine its confluence. For adherent cells, which grow attached to a surface, passaging is performed when they reach 70% to 90% confluence, meaning they cover that percentage of the vessel’s surface. Suspension cells, which grow floating in the medium, are passaged when they begin to clump or the medium appears turbid.
Once ready, adherent cells need to be detached from their growth surface. This is commonly achieved through enzymatic treatment, using trypsin, a proteolytic enzyme that cleaves proteins responsible for cell adhesion. Trypsin solutions often include EDTA, which further weakens cell interactions and enhances detachment. Alternatively, mechanical scraping using a cell scraper can be employed to detach cells, especially for sensitive cell types.
After detachment, the cells are collected, often by centrifugation, to form a pellet. This cell pellet is then resuspended in a fresh growth medium, and the cells are counted to determine their concentration and viability. Based on the desired seeding density for the new culture vessels, the cell suspension is diluted and transferred into fresh flasks or dishes containing new growth medium. Maintaining a sterile environment throughout this entire process, known as aseptic technique, is essential to prevent microbial contamination.
The Role of Cell Passaging in Research and Medicine
Maintaining healthy, growing cell lines through routine passaging is important for numerous scientific and medical advancements. In drug discovery, passaged cells serve as models to test the efficacy and toxicity of new chemical compounds before they proceed to animal and clinical trials. This allows researchers to screen potential drug candidates and understand their pharmacological actions on human cells.
Cell passaging also plays a role in vaccine production, where viruses are grown in cell cultures to create viral vaccines for diseases such as influenza, measles, and polio. Live attenuated vaccines are developed by serially passaging wild-type viral pathogens in cell cultures, which can lead to a weakened, less virulent strain suitable for vaccination.
In disease modeling, passaged cell lines are used to create in vitro representations of human diseases, such as cancer or neurodegenerative disorders. By culturing cells from patients or genetically modifying cell lines, scientists can investigate the cellular and molecular mechanisms of diseases and test potential therapeutic interventions. Passaging is also important in regenerative medicine and tissue engineering, where cells are grown and expanded for therapeutic purposes, such as creating skin grafts for burn patients or developing artificial tissues for organ repair.