Hi All,
I am now using the lab-tek chamber slide to do the cell biology experiment. But I found it is very hard to seed the cell very evenly. Cell seeding is important for me as my final readout is cell-density dependent. Can you guys give me some suggestions on that? Thanks a lot.
Gang
Try perpendicular shaking before you put your cells in the incubator. Any rotational movement will result in accumulation of your cells in the central region of the well chambers.
Hope that information helps!
Best regards,
Christian
Hi! When you completed the cell seeding into the chambers on the slide, just leave it idle for 10-20 min, so the cells would sediment and start anchoring. Then transfer the slides into the CO2 incubator accurately. Good luck!
Try perpendicular shaking before you put your cells in the incubator. Any rotational movement will result in accumulation of your cells in the central region of the well chambers.
Hope that information helps!
Best regards,
Christian
Hi. There are several options to do that.
First of all you should control the number of cells. If there are too much of them - they will be confluent for the next day. Usually I add 30-50k cells in each well of two-well Labtek chamber
The second the pipetting is our everything. However, you should spread the cells for the whole bottom of the chamber and do that gently. any kinds of shaking usually result in concentration of the cells in center of the chamber.
The third, if your experiment allows, you may wash your cells with medium after 5-10 hours after seeding. For me it works perfectly fine. In this way the weak and confluent cells will be washed.
One important thing. It is better to add the low volume of cells in medium to the chamber, just to cover the bottom and then add the medium. I this way the cells will faster get the bottom of the chamber and the distribution will be more homogenous. Here you do not need pipetting.
Also you may control the cell growth with the medium complemented with different percent of serum.
Best regards,
Kyrylo
What would be final volume of media per well? What coating system I can use for Neonatal rat cardiomyocyte cell seeding? Thank you
Dont know coating for other cells but neurons and mesenchymal cells work fine in LabTekII with Poly-D-lysine 0.01% (final conc.). Seeding: 25.000 cels/well , 0.2-0.5ml. You should check them (it depends of your cell type) since a minor surface could increase their growth in confluency. Usually they grow easily on corners. (Also,..totally agree with Dr Christian Wechselberger comment -above-).
Hi There I want to seed cardiac fibroblasts in Chamber slide Lab-TeK II 154526. Anyone could share with me his protocol. Thanks.
i am using 8 well labteks, and with seeding them my problem was NOT concentration in the center but on the corners. maybe because its too small.
cell density is according to your cells and their doubling time .however i suggest the method said before about adding in small volume at first then adding medium after 10 to 20 mins . gentle shaking is good if you are facing the same problem as me (center emptiness) or if you are using 8 well labteks (i use millicell ones )
also , merci for your suggestions too i might just mix them up to get better results!
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