User:Wilfred J. Poppinga/Notebook/cAMP compartmentalization/2010/03/17

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Ht31P toxicity Main project page
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Summary

  • Stop stimulation
  • Prepare washing buffer 1x ELISA + 10x PBS
  • CoIP

Materials & Methods

Method

Preparations for ELISA & Viability staining

Co-immunoprecipitation Part III

  • Centrifuge sample incubated with beads, 5 min. 14.000 RPM
  • Remove supernatant* (w. syringe)
  • Add 1 mL PBS
  • Centrifuge, 5 min. 14.000 RPM
  • Remove supernatant*
  • Add 75 μL Laemli buffer to beads
  • Store samples @ -20 °C


* Be careful not to touch beads

Results

  • PICTURES OF CELL DEATH (Anouk)
  • Viability staining
Ht31P (μM) Ht31P (μM) Ht31P (μM) Ht31P (μM) Ht31P (μM) Ht31P (μM)
D9 P23 0 10 20 30 40 50
CTR 752441172092192913461712
CTR 671930301540132311261458
CSE 3996922649547527660
CSE 35091125646608550577

Image:SurvivalRateD9P2317032010.JPG

Ht31P (μM) Ht31P (μM) Ht31P (μM) Ht31P (μM) Ht31P (μM) Ht31P (μM)
D12 P27 0 10 20 30 40 50
CTR 715152423640295324352331
CTR 681643532746233919702074
CSE 528417841234939938918
CSE 4858189112501020870907

Image:SurvivalRateD12P2717032010.JPG

Conclusion

  • Ht31P is lethal for airway smooth muscle hTERT cells
  • Strength of effect is donor dependent



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