Western Blot Imaging Tutorial

List of Items needed:
-Membranes
-Imaging Solution kit
- Flat nosed tweezers
-Parafilm
-Plastic Wrap
-1 ml pipet
-pipet tips
-1.5 ml Eppendorf tubes
-USB drive
-Paper towels
-garbage bin (optional)
Notes:
It is critical that you do not mix stock solution A and B. Remember to change tips before going from one stock solution to the next.
Once the solution is mixed, it is slightly light sensitive. If, for example, you wanted to make two 1.5 ml tubes of imaging solution before using them to save time, keep one of them in the shadow of one of the machines or underneath something so that it is less exposed to light.
Only make the amount of imaging solution you know you will use. I use 800 ul of prepared solution (which is solution A mixed with B) per half a membrane.
If I have more membranes that I need to image after the first, than I don't discard the solution that is on top of the plastic wrap that I used for the first. I continue to use that same solution but add another 400 ul of fresh solution to what is already there.
I continue this process for about 4 half-membranes and then I would start with a fresh piece of plastic wrap, discarding the old solution and plastic wrap and getting a new one.
Ensure that the face of the membrane makes good contact with the imaging solution. It is okay to gently touch the back side of the membrane to ensure proper contact.
Avoid air bubbles when laying the membrane down onto the parafilm in the imaging machine. Sometimes you don't see the small air bubbles until you image which can be frustrating if you don't know what to look for. Instead of placing the membrane down as quickly as possible onto the parafilm, place one edge down first and slowly lie it down so that any air bubbles that are present may move to the opposite edge.
The membrane can also be physically moved around on the parafilm so long as you are only touching the edges.
Try to keep the edges of the membrane parallel to the edges of the parafilm so it looks nice.
If the bands look really weak initially, it is okay to put the membrane back into the imaging solution and use even more if you think it is necessary.
Remember that imaging the membrane is more of an art skill than a technical skill, you'll have to play around with the amount of solution you use, how long you keep the membrane in the imaging solution, how you lay it down on the parafilm, and what imaging exposure settings (in the machine) you use to get a good image. It will take some practice but it's not much different than taking a picture with a camera once you're used to it.
Please remember to clean up after use