Hosuk:LabNotes/2013-10-28

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Revision as of 20:44, 28 October 2013 by >Hosuki78 (→‎Procedure)
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Detect RAB7A, and compare to ACTB

  • Use 10/22 sample (MatTek dish, S2)
  • RAB7A : Resuspension --> 200uM
  • 1uL 200uM RAB7A + 2uL Cy5 1st rolony adapter in 197 uL 2x SSC


Preparation

  • 1uL 200uM ACTB + 199 uL 2x SSC --> 2 tubes
  • 1uL 200uM RAB7A + 199 uL 2x SSC --> 2 tubes
  • 1uL 200uM ACTB + 2uL Cy5 1st rolony adapter in 197 uL 2x SSC --> 2 tubes
  • 1uL 200uM RAB7A + 2uL Cy5 1st rolony adapter in 197 uL 2x SSC --> 2 tubes
  • 2mL 80% Formamide
  • 2x SSC, PBS
  • Aspiration setup, vacuum pump


Procedure

  • Strip, Add 100uL (RAB7A+1st rolony probe mix), image --> 1st round RAB7A
  • Strip, Add 100uL (ACTB+1st rolony probe mix), image --> 1st round ACTB
  • Strip, Add 100uL (RAB7A+1st rolony probe mix), image --> 2nd round RAB7A
  • Strip, Add 100uL (ACTB+1st rolony probe mix), image --> 2nd round ACTB
  • Strip, Add 100uL (RAB7A only), image --> 1st round RAB7A only
  • Strip, Add 100uL (ACTB only), image --> 1st round ACTB only
  • Strip, Add 100uL (RAB7A only), image --> 2nd round RAB7A only
  • Strip, Add 100uL (ACTB only), image --> 2nd round ACTB only