Cell synchronization

Cell Synchronization. Synchronization literally means to make two or more things happen exactly simultaneously. For instance, two or more watches can be synchronized to show exactly the same time. The cells at different stages of the cell cycle in a culture can be synchronized so that the cells will be at the same phase. Cell synchrony is required to study the progression of cells through cell cycle. They are broadly categorized into two groups - Physical Fractionation for Cell Separation and Chemical Blockade for Cell Separation.

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Cell separation by physical means

Physical fractionation or cell separation techniques, based on the following characteristics are in use.

• Cell Density
• Cell Size
• Affinity of Antibodies on cell surface epitopes.
• Light Scatter or fluorescent emission by labelled cells.

The two commonly used techniques are:

Centrifugal Separation

The physical characteristics- cell size and sedimentation velocity are operative in the technique of centrifugal elutriation. Centrifugal elutriator (from Beckman) is an advanced device for increasing the sedimentation rate so that the yield and resolution of cells is better. The cell separation is carried out in a specially designed centrifuge and rotor.

Fluorescence-activated Cell Sorting

Fluorescence-activated cell sorting is a technique for sorting out the cells based on the differences that can be detected by light scatter (eg. cell size) or fluorescence emission (by penetrated DNA, RNA, proteins, antigens). The procedure involves passing of a single stream of cells through a laser beam so that the scattered light from the cells can be detected and recorded. There are two instruments in use based on its principle:

• a) Flow cytometer
• b) Fluorescent-Activated Cell Sorter (FACS)

Cell separation by chemical blockade

The cells can be separated by blocking metabolic reactions. Two types of metabolic blockades are in use:

Inhibition of DNA Synthesis

During the S Phase of cell cycle, DNA synthesis can be inhibited by using inhibitors such as thymidine, aminopterin, hydroxyurea and cytosine arabinoside. The effects of these inhibitors are variable. The cell cycle is predominantly blocked in S phase that results in Viable Cells.

Nutritional Deprivation

Elimination of serum from the culture medium for about 24 hours results in the accumulation of cells at G1 phase. This effect of nutritional deprivation can be restored by their addition by which time the cell synchrony occurs.