New year and already things are off and running.
You know how some days you have one of those head-slap moments — a “how did I not think of that before” moments.
This week, the first of the year, I had one of those moments.
A few years ago I had been cranking out Western blots (a technique to look at proteins)* and all was going well. Then there was a three-year gap because I was working in another lab. Upon my return to my “old” lab, I started up the ol’ Western blotting operation again — only this time things did not go so smoothly.
The numerous protein bands I had come to expect on the blots just weren’t there. Increasing the amount of protein I added to each sample didn’t help. Making new buffers and new formulations of those buffers didn’t help. Using new reagents to detect those proteins didn’t help.
It wasn’t until I ran out of the nitrocellulose membrane – the paper-like substance the proteins were stuck on – and I opened a new box of membrane that it became obvious what the problem was. The membrane stock I had been using was over four or five years old. Things had happened to that membrane – things like exposure to air and temperature fluctuations – caused the little charges that helped hold the proteins onto the membrane to change in some way and loose their ability to grip the proteins.
This was something I so obviously should have considered when I was trying to figure out why the procedure wasn’t working like it had a few years before.
So, as you can see by the photos I’ve included in this post, there are more protein bands and they are “crispier” on the new membrane than on the old membrane.
I guess, after 20 years in the lab, I still have a few things left to learn.
*If you would like to see how a Western blot is done, see my previous post.