反应详情

The survival of transfected cells in the absence or presence of G418 was monitored by staining of foci/clones with neutral red in vivo with subsequent crystal violet staining. The medium was removed from the cells and replaced with fresh medium containing 0.0025% neutral red (Sigma N2889) and the cells incubated for 3 hours at 37° C. Cells were washed twice with PBS, fixed in 3.5% formaldehyde for 15 minutes, washed twice again in PBS and then with distilled water and the number of (live) foci counted. The cells could then be re-stained with crystal violet by incubating with an 0.1% crystal violet (Sigma C0775) solution in 20% methanol for 20 minutes at room temperature, followed by 3 washes in 20% methanol and a wash with distilled water.

1. customThe medium was removed from the cells
2. additioncontaining 0.0025% neutral red (Sigma N2889)
3. washCells were washed twice with PBS
4. waitfixed in 3.5% formaldehyde for 15 minutes
5. washwashed twice again in PBS
6. waitby incubating with an 0.1% crystal violet (Sigma C0775) solution in 20% methanol for 20 minutes at room temperature
7. washin 20% methanol and a wash with distilled water