The best alternative iPS Cells department description:
Department of iPS CellsTechnical director Dr Goffer worked at CiRA research institute in Japan, iPS Cells with many years of rich production experience, lead the department of technical personnel by Japan's Kyoto university standard technology, combined with the first national patent technology making iPS Cells, simple operation, stick a wall high success rate, cell is stable, and use the patent transportation container transportation iPS Cells, to better ensure the quality of cells.Company also opened a dedicated iPS Cells technology practice and training services, can be customized according to customer's requirements practice content, details see the web site"Technology practice"Page, or call advisory telephone 400-021-2021.
The cell description:
Induced human foreskin cells into IPS Cells by reprogramming transcription factors as: OCT4, SOX2, KLF4, MYC induced to establish IPS Cells, culture without feeding layer cells.
IPS training procedures
1, in advance with the Matrix Petri dish / board coating processing. Usually Matrix stored at -20 ℃, before use on the refrigerator at 4 ℃ or ice on the thaw. To be Matrix thawed, according to the ratio of 1: 100 quickly diluted with PBS liquid. Add 1ml, add 6ml dish and add 2ml, 10cm dish and add 3ml of the amount of added, after adding quickly shake the plate / board, so that the Matrix completely cover the entire bottom of the board. Into the 37 ℃ incubator for 4 hours, before use to remove the coating solution (if not used, sealed with a sealed membrane, refrigerator at 4 ℃ can save a week).
2, prepare the PSCeasy Thawed Complete Medium (PSCeasy Thaw Base and PSCeasy Thaw Additive) before resuscitation. Add the appropriate amount of thawing solution (6-well plate for each hole plus 2ml, 6cm dish plus 3ml, 10cm dish plus 9ml), add another 10ug / ml Y27632 factor, into the 37 ℃ incubator.
3, the recovery of a cell with 5ml of the above mixed medium, before the recovery in the 37 ℃ water bath full preheat. Remove the cryopreservation tube from the liquid nitrogen tank and quickly transfer it to the biological safety cabinet (if the position of the liquid nitrogen tank is far from the safety cabinet, transfer the cryopreservation tube to the safety cabinet with a liquid nitrogen container) Good complete medium for thawing (with the pipettes kept into the cryopreserved into the extraction medium, exchange the dissolved cells until completely dissolved).
4,200Xg centrifuge for 5 minutes, remove the supernatant, add 1ml of PSCeasy completely thawed medium (containing 10ug / ml Y27632 factor), with the fingers of the tube tube collapse. Add the PSCeasy completely to the centrifuge tube by inoculating the amount of each plate / dish 1 ml, gently aspirating three or four times into the dish / plate. Horizontal cross vibration to cells evenly distributed, 5% CO2, 37 ℃ incubator for 24 hours.
5,After 5 hours, replace PSCeasy complete medium (PSCeasy basal medium and PSCeasy basic medium additive). After every 22-24 hours for liquid. Cells covered with 80-90% need to pass or freeze.
Second, pass / frozen
1, before passing the petri dish / board coating treatment, practice and recovery the same. The medium was PSCeasy complete medium, plus 10 μl / ml Y27632 factor.
2, before the pass / cryopreservation, prepared 37 ℃ preheated good good calcium and magnesium solution and the passage of the solution (0.5mM EDTA + 0.025% trypsin).
3, aspirate PSCeasy complete medium, and add 37 ℃ preheated PBS solution to wash twice.
4, add appropriate amount (by 6-well plate plus 1ml per hole, 6cm dish plus 2ml, 10cm dish plus 3ml) of 37 ℃ preheated good passage of working fluid.
5,37 ° C for 4-5 minutes (after a minute later, with the fingers flap several plates / bottom of the dish), observed under the microscope most of the cloned edge began to escape from the plate / plate bottom, clone most of the internal Cell gap between the cells have not yet separated from each other, the naked eye observed cell colonies become opaque and white, indicating that the cell digestion time is ideal.
6, quickly absorb the passage of liquid, add the appropriate amount of PSCeasy complete medium to terminate the digestion, with a pipette fan-shaped dish / plate bottom (no more than 10 times), so that the attached cell colonies off. (If the digestion time is improper, the cells are completely peeled off from the plate / plate, without the addition of the passage, add the same amount of PSCeasy complete medium to terminate the digestion).
7, into the centrifuge tube, after the centrifugal resuspend. Pass the ratio of 1: 8-1: 12; frozen by 6-well plate per hole 1, 6cm frozen 2-4, 10cm frozen 6-12. Each add 0.8 ml of 90% PSCeasy complete medium with 10% DMSO.
8, the recovery by the pre-frozen 1: 4-1: 6 ratio.
|Serial number||product name||Item number||specification||Price|
|Two||Ips work transfer fluid||500ml||210.00||icell-CA3001500|
|Three||Ips basic culture medium||500ml||1785.00||icell-CA1001500|
|Four||Defrost liquid ips||100ml||738.00||icell-CA1002100|
1. iPS Cells operation instructions .pdf download