Daniel:Notebook/RNAFISH/2014-9-3: Difference between revisions
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>Djacobse (Created page with "=Trial Run (Started 08/29/2014)= Back to Calendar ==Post-Hybidization== #Remove rubber cement together wit...") |
>Djacobse |
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#Remove rubber cement together with the chamber using a pair of fine forceps | #Remove rubber cement together with the chamber using a pair of fine forceps | ||
#Wash slides in | #Wash slides in 2X SSC at 37°C, 3 × 15 min (preferably with shaking) to remove hybridization mixture | ||
#Perform stringency washes in 0.1X SSC at 60°C, 2 × 5 min to remove nonspecific hybrids | #Perform stringency washes in 0.1X SSC at 60°C, 2 × 5 min to remove nonspecific hybrids | ||
#Equilibrate in 2X SSC for 2 min | #Equilibrate in 2X SSC for 2 min |
Revision as of 16:02, 3 September 2014
Trial Run (Started 08/29/2014)
Post-Hybidization
- Remove rubber cement together with the chamber using a pair of fine forceps
- Wash slides in 2X SSC at 37°C, 3 × 15 min (preferably with shaking) to remove hybridization mixture
- Perform stringency washes in 0.1X SSC at 60°C, 2 × 5 min to remove nonspecific hybrids
- Equilibrate in 2X SSC for 2 min
- Add DAPI, let sit for at least 30 minutes
- Seal the coverslip with nail polish; let sit 30 minutes
- Sections become fragile after the FISH; therefore, strong pressure on the coverslip should be avoided
- e.g., pressing the coverslip with the microscope objective when focusing often completely destroys the sample