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Team:Penn/Notebook
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<p>First time doing calcium imaging. Brought over 293T's in a 6 well plate, which had been transfected with CatCh. Also had positive control cells on which we used ionophores. Used the Meaney Lab confocal and x-rhod1 dye. Media had 2mM Calcium. | <p>First time doing calcium imaging. Brought over 293T's in a 6 well plate, which had been transfected with CatCh. Also had positive control cells on which we used ionophores. Used the Meaney Lab confocal and x-rhod1 dye. Media had 2mM Calcium. | ||
1uM Ionomycin addition did not change x-rhod1 fluorescent intensity in Anthony's neurons. | 1uM Ionomycin addition did not change x-rhod1 fluorescent intensity in Anthony's neurons. | ||
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Also did not work on our 293T's, but we had the problem of being unable to cleanly pipet the ionomycin into the 6 well plates because the wells we looked at with the scope became entirely blocked by the scope lens. | Also did not work on our 293T's, but we had the problem of being unable to cleanly pipet the ionomycin into the 6 well plates because the wells we looked at with the scope became entirely blocked by the scope lens. | ||
Illumination of CatCh cells with 476nm laser from confocal did not increase x-rhod1 intensity. eYFP expressing cells were identified, so transfection worked. | Illumination of CatCh cells with 476nm laser from confocal did not increase x-rhod1 intensity. eYFP expressing cells were identified, so transfection worked. | ||
Latest revision as of 02:11, 25 September 2011
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Notebook
09/20/2011 (Avin, Mike M, Peter, Anthony)
First time doing calcium imaging. Brought over 293T's in a 6 well plate, which had been transfected with CatCh. Also had positive control cells on which we used ionophores. Used the Meaney Lab confocal and x-rhod1 dye. Media had 2mM Calcium.
1uM Ionomycin addition did not change x-rhod1 fluorescent intensity in Anthony's neurons.
Also did not work on our 293T's, but we had the problem of being unable to cleanly pipet the ionomycin into the 6 well plates because the wells we looked at with the scope became entirely blocked by the scope lens.
Illumination of CatCh cells with 476nm laser from confocal did not increase x-rhod1 intensity. eYFP expressing cells were identified, so transfection worked.
Will try again Thursday and Friday with larger plates, and possibly switch to FURA-2 and Avin's laser if the confocal still doesn't work. Anthony is going to try further with x-rhod1 dye in neurons to make sure that the dye/laser parameters aren't the problem. Will also attempt the co-culture experiment on these dates.
2011 Penn iGEM Team.
