Difference between revisions of "Developing Your Immunoblot Membrane"

From SDMRC
Jump to: navigation, search
(Created page with "==Developing your Immunoblot Membrane== ===Materials=== *A flash drive *A tube rack *Sheet protectors *PICO developing solution *15 ml tube *Ethanol + KimWipes towels # Afte...")
 
 
Line 1: Line 1:
==Developing your Immunoblot Membrane==
 
 
===Materials===
 
===Materials===
 
*A flash drive  
 
*A flash drive  
Line 7: Line 6:
 
*15 ml tube
 
*15 ml tube
 
*Ethanol + KimWipes towels
 
*Ethanol + KimWipes towels
 +
 +
===Method===
 
# After the 1 hour secondary time has passed take your membranes into trays of TBST to wash three times for 10 minutes   
 
# After the 1 hour secondary time has passed take your membranes into trays of TBST to wash three times for 10 minutes   
 
# Cut new sheet protector for your membranes (they can be in one protector)   
 
# Cut new sheet protector for your membranes (they can be in one protector)   

Latest revision as of 20:18, 23 July 2018

Materials

  • A flash drive
  • A tube rack
  • Sheet protectors
  • PICO developing solution
  • 15 ml tube
  • Ethanol + KimWipes towels

Method

  1. After the 1 hour secondary time has passed take your membranes into trays of TBST to wash three times for 10 minutes
  2. Cut new sheet protector for your membranes (they can be in one protector)
  3. Take your pico developing solutions (two bottles on bench: brown and clear) with a 15 ml tube, a pipette, and a tube rack to the scanner
  4. Quickly take your membranes into the plastic (dry a little) and bring to the scanner
  5. AT THE SCANNER mix the brown and white solution in the 15 ml tube and flush
  6. The developing solution is 1:1 so divide the total amount needed by 2

Example: 4.5 ml total so 2.25 ml of brown and 2.25 ml of clear

Developing Machine

  1. Open the tray and clean with ethanol
  2. Open the sheet protector and quickly add drops of the developing solution until is covers the membranes
  3. Leave the sheet open and push tray in
  4. On the scanner hit live view to see your membranes and adjust position if needed
  5. Hit application_chemiluminesence_ blot_exposure_ show preview optimal auto exposure
  6. Place red crop box over the area you want captured and hit camera and wait
  7. (optional) manual exposure_ set manual exposure time (ex. 60 sec) _ capture and wait. Make sure you are not moving the membrane during these pictures.
  8. Take a full picture at the end. Application_ colormetric_ exposure (automatically/ faint bands) _ capture
  9. Go to gallery_ take any picture and the one you just took (select both)_ hit merge_ back to gallery
  10. Put all pictures wanted onto the flash drive_ save/send_ save to USB stick
  11. Trash membranes, clean tray with ethanol

Add the ponceau scan to your other developed images

  • Copy a paste ponceau scan to your flashdrive
  • Make new folder, title with condition and cell type
  • Put all the images from flash drive onto personal computer