Frequently Asked Questions about Cell Lines

Chosing a line

Growing cells

Immunohistochemistry

Which line should I use?

If you are looking for a general all-purpose cell line to use for biochemistry or for ectopic expression of your favorite protein, you should use either Kc167 or S2-DGRC, both of which are easy to grow and have been widely used for many years. S2R+ is an isolate of S2 which is used by many labs because it spreads on the substrate and is therefore more accessible for morphological studies. S2-DRSC is the isolate of S2 that is used routinely for RNAi experiments by the DRSC.

If you are looking for particular cell properties, you may find the data generated by modENCODE to be helpful. modENCODE is collecting extensive data on 4 cell lines (Kc167, S2-DRSC, ML-DmBG3-c2, and CME-W1-Cl.8+), and the fly transcriptome group within modENCODE is collecting data on 21 additional lines. These data are available from the modENCODE web-site; data will continue to be added to this site at least through 2011.

The lines in our collection probably represent a wide variety of cell types, but they probably are not exact duplicates of the cell types from which they are derived. Transcriptome data indicates a wide range of cell types among the lines derived from a single tissue source. In the end, you will have to test for yourself to see whether the pathway you are interested in is preserved in any given line.

The cells aren't growing. What should I do?

If you ordered Kc or S2 or S2R+, you are doing something wrong. But most of the disc and CNS lines are slow growing and slow to recover from freezing. They should be treated very gently (don't jostle any more than necessary, and don't change the medium all at once). Be patient; it may take weeks or even months for them to take off. It generally helps to replace about half the medium with fresh medium every week or so. If you cannot get the cells to grow, contact us at dgrc@cgb.indiana.edu; we can often help with trouble-shooting, and if it is necessary, we may try to thaw them at the DGRC and send growing cells to you.

What medium should I use?

The recommended medium is listed on the web-page for each cell line. We use M3+BPYE for most lines, which we make in-house. There is a recipe in the cell line protocols on our web-site. This medium is very similar to Schneider's medium, and you can substitute Schneider's medium for M3+BPYE for many of our lines. We have not tested Schneider's medium on most of our lines, and in some cases, we have found that the cells do not grow as well in Schneider's medium as in M3+BPYE, so you should use Schneider's medium with caution. All cell lines require addition of fetal calf serum to both M3+BPYE and to Schneider's medium, and individual cell lines require one or more other additions to this medium (insulin, fly extract, selective agents); these are described in detail on our website. Some lines need M3 medium rather than M3+BPYE, and we routinely grow Kc167 in CCM3 (without the addition of serum), though it can also be grown in M3+BPYE with serum.

Should I include antibiotics in the medium?

We don't use antibiotics; if you have good sterile technique, they should be unnecessary. On the other hand, they probably don't do any harm. If you wish to use antibiotics, try pen/strep at the concentration used for mammalian cells.

I just thawed my cells, and the culture is contaminated. What should I do?

Unless we have reason to think that our stock is contaminated, we usually ask you to re-order. But please tell us, because user feed-back helps us know know if there is a problem with the stock.

Where can I get fly extract?

You can make it yourself, using this protocol or you can purchase it from the DGRC.

What size of poly-L lysine should I use?

We recommend purchasing lyophilized poly-L lysine from Sigma with a molecular weight of 300,000 or larger. 1% or 10% solutions can be made up and stored at -20. The working concentration is 1% and can be stored for up to 3 months at 4 degrees. We typically dip slides in the 1% solution and reuse the solution several times.


Citing the DGRC

When publishing experiments using materials obtained from the DGRC please cite the Drosophila Genomics Resource Center, supported by NIH grant 2P40OD010949, in the acknowledgments. Your cooperation helps us when we need to renew our grant.