Agarose is a naturally occurring material derived from specific types of red algae. It is a fundamental component in many laboratory procedures, recognized for its unique properties that enable delicate separations and specialized environments for biological studies.
Defining Agarose
Agarose is a linear polysaccharide primarily extracted from certain red algae species, such as Gelidium and Gracilaria. It consists of repeating disaccharide units called agarobiose, composed of D-galactose and 3,6-anhydro-L-galactopyranose. This forms a long, unbranched polymer chain.
Agarose is distinct from agar, its crude precursor. Agar is a mixture containing both agarose and agaropectin. Agaropectin contains charged groups like sulfates and pyruvate, making agar less suitable for certain applications. Agarose is the purified, neutral component of agar, responsible for its gelling properties and preferred for high-purity applications.
The Unique Characteristics of Agarose Gels
Agarose’s utility stems from its ability to form a stable, porous gel. Gelation occurs when agarose powder is dissolved in hot water or buffer and then cooled. As the solution cools, linear agarose polymer chains associate non-covalently through hydrogen bonds, forming a three-dimensional network of helical fibers. This network traps water molecules, creating a semi-solid matrix.
Agarose gels have adjustable pore sizes. The concentration of agarose directly influences the size of these microscopic pores; higher concentrations result in smaller pores, while lower concentrations yield larger pores. This property allows researchers to tailor the gel to separate molecules of different sizes. Agarose is largely inert, meaning it does not chemically interact with most biological molecules. This inertness and optical clarity make it an excellent medium for observing and separating delicate biomolecules.
Primary Uses of Agarose in Science
Agarose is widely used in gel electrophoresis, particularly for separating DNA and RNA fragments. In this technique, an electric field causes negatively charged nucleic acids to migrate through the agarose gel towards the positive electrode. The gel acts as a molecular sieve; smaller fragments navigate through the pores more quickly than larger ones, allowing separation based on size. Agarose is favored for separating DNA fragments ranging from approximately 100 base pairs to 25 kilobases, and even larger fragments up to several megabases with specialized techniques. It is often preferred over polyacrylamide gels for larger molecules due to its larger pore size and ease of preparation.
Agarose is also used in chromatography methods, such as gel filtration. Here, agarose beads, often cross-linked for stability, separate molecules by size as they pass through a column. Larger molecules pass through more quickly, excluded from the beads’ pores, while smaller molecules enter the pores and take a longer, more tortuous path.
Beyond separations, agarose is used in cell culture to create three-dimensional (3D) environments for cell growth. Cells can be cultured within or on agarose matrices, which provide a non-adhesive surface encouraging spheroid or aggregate formation, mimicking natural tissue structures. This is useful in cancer research, such as the soft agar assay, where transformed cells’ ability to grow independently in a semi-solid agarose medium is assessed. The porous nature of agarose allows for nutrient diffusion and gas exchange, supporting cell viability in these 3D models.