In the researches of living cells, especially the cells with pluripotency to differentiate such as iPS cells, ES cells and other stem cells, it is essential to use single cells of good quality in order to obtain the cell population (cell colony) with the same genetic background and biological characters for further study.
It is known that human iPS cells have a susceptibility to physical damage, and they undergo apoptosis with a high rate after passaging of single cells. Although the cell survival rate has been improved following the application of the Rho-associated kinase (ROCK) inhibitor, passaging is typically performed in the state of uneven small masses of cells (clumps). To exactly prove the property of pluripotency of stem cells, it is essential to utilize single cell-derived (monoclonal) cells. In case of iPS cells established by colony pickups, however, their homogenicity is not guaranteed.
Although conventional high-speed droplet-charging sorting technique using ultrasonic waves, high-voltage charging and high hydraulic pressures can efficiently perform analysis and sorting, there is concern that these sorting methods may cause considerable physical damage and have an unneglected affection on living cells.
Our PERFLOW Sort does not use ultrasonic waves or high-voltage charging, and besides, it manipulates liquid with a water pressure that is less than one tenth of conventional systems. Moreover, PERFLOW Sort has a short and straight pipe conduit for the transportation of cells in order to remove the second flow of fluid. As a result, the shear force for living cells can be eliminated. Thus, the non-droplet cell sorting method of PERFLOW Sort decreases to the utmost limit the main physical damage factors in the sorting process.
Importance of damage-less sorting
- Differentiation induction in iPS cells, ES cells and other stem cells
- Decreased risk of malignant transformation resulted from genetic instability
- Proper evaluation of biological behaviors in the researches of gene engineering and drug development
- Acquisition of a large number of single cell-derived stem cells
Non-droplet cell sorting
In non-droplet cell sorting, the flow velocity of target cells is accurately measured by the transmitted light measuring system with optical fibers aligned in the cell flow, based on which the arrival time of the target cell to the nozzle end is calculated, and mechanical changeover is appropriately manipulated between waste tank and culture plate.
The non-droplet cell sorting method enables damage-less sorting of delicate living cells independently of their size.
Mechanism of Damage-less Sorting
Comparison of sorting conditions between conventional sorters and the PERFLOW series
|Damage-causing factors||Droplet-charge Method
|Supersonic wave (kHz)||20～200||0|
|Shock kinetic energy(Note 1)||>1,500||1|
|Shear power by fluid(Note 2)||+||- (vertical tube)|
(Note 1):Comparison of kinetic energy(J).
Shock kinetic energy was calculated using that received by cells at the time of touching on water by the No-drop-no-charge method as 1 (calculated assuming a drop velocity of 10m/sec.). Back to Main Content
(Note 2):Shear power by the fluid refers to the shear power the cells received when they pass through the flow with flexion, where secondary flow occurs and causes cells to move in a spiral course. Back to Main Content
The main physical damage factors have been decreased to the utmost limit in the sorting process of PERFLOW sort, therefore achieving damage-less sorting of living cells.
How Damage-less cell sorting works
A video showing how cell sorting works is available.