Cell synchronization
Encyclopedia
Cell Synchronization is a process by which cells at different stages of the cell cycle
in a culture are brought to the same phase. "Cell synchrony" is required to study the progression of cells through the cell cycle. The types of synchronizations are broadly categorized into two groups: "Physical Fractionation" and "Chemical Blockade."
The two commonly used techniques are:
. 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.
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:
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.
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.
Cell cycle
The cell cycle, or cell-division cycle, is the series of events that takes place in a cell leading to its division and duplication . In cells without a nucleus , the cell cycle occurs via a process termed binary fission...
in a culture are brought to the same phase. "Cell synchrony" is required to study the progression of cells through the cell cycle. The types of synchronizations are broadly categorized into two groups: "Physical Fractionation" and "Chemical Blockade."
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 epitopeEpitopeAn epitope, also known as antigenic determinant, is the part of an antigen that is recognized by the immune system, specifically by antibodies, B cells, or T cells. The part of an antibody that recognizes the epitope is called a paratope...
s. - Light scatter or fluorescent emission by labeled cells.
The two commonly used techniques are:
Centrifugal separation
The physical characteristics — cell size and sedimentation velocity — are operative in the technique of centrifugal elutriationElutriation
Elutriation, also known as air classification, is a process for separating lighter particles from heavier ones using a vertically-directed stream of gas or liquid . This method is predominately used for particles with size...
. 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 (FACS) is a technique for sorting out the cells based on the differences that can be detected by light scatter (e.g. cell size) or fluorescence emission (by penetrated DNA, RNA, proteins, antigens).The procedure involves passing of a single stream of cells through a laser
Laser
A laser is a device that emits light through a process of optical amplification based on the stimulated emission of photons. The term "laser" originated as an acronym for Light Amplification by Stimulated Emission of Radiation...
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) Fluorescence-activated cell sorter
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 phaseS phase
S-phase is the part of the cell cycle in which DNA is replicated, occurring between G1 phase and G2 phase. Precise and accurate DNA replication is necessary to prevent genetic abnormalities which often lead to cell death or disease. Due to the importance, the regulatory pathways that govern this...
of cell cycle, DNA synthesis can be inhibited by using inhibitors such as thymidine
Thymidine
Thymidine is a chemical compound, more precisely a pyrimidine deoxynucleoside. Deoxythymidine is the DNA nucleoside T, which pairs with deoxyadenosine in double-stranded DNA...
, aminopterin
Aminopterin
Aminopterin , a 4-amino analog of folic acid, is an antineoplastic drug with immunosuppressive properties used in chemotherapy. Aminopterin is a synthetic derivative of pterin. Aminopterin works as an enzyme inhibitor by competing for the folate binding site of the enzyme dihydrofolate reductase...
, hydroxyurea
Hydroxyurea
Hydroxycarbamide or hydroxyurea is an antineoplastic drug, first synthesized in 1869, used in myeloproliferative disorders, specifically polycythemia vera and essential thrombocythemia...
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 serumBlood plasma
Blood plasma is the straw-colored liquid component of blood in which the blood cells in whole blood are normally suspended. It makes up about 55% of the total blood volume. It is the intravascular fluid part of extracellular fluid...
from the culture medium for about 24 hours results in the accumulation of cells at G1 phase
G1 phase
The G1 phase is a period in the cell cycle during interphase, before the S phase. For many cells, this phase is the major period of cell growth during its lifespan. During this stage new organelles are being synthesized, so the cell requires both structural proteins and enzymes, resulting in great...
. This effect of nutritional deprivation can be restored by their addition by which time the cell synchrony occurs.