Debbie Knight

A Day in the Life: March 7, 2013

In A Day in the Life, research log on March 7, 2013 at 4:56 pm

From time to time, I will give a glimpse into the “glamorous” life of a research associate and talk about what I’m doing in the lab. These entries I will call “A Day in the Life…” 

Today I’m searching for an antibody that I can use to stain the cells that line blood vessels (endothelial cells) in mouse tissue.

And I want to visualize where the antibody binds in that tissue with a dye that glows (or fluoresces) under certain conditions.  Something like this:

MEK14.7 ab8158_3

From abcam.com: Ab8158 at a dilution of 1/50 staining CD34 from mouse lung cells by Immunocytochemistry

A Google search has turned up several results that look promising.

However, there’s some cross checking that has to happen before my lab will plunk down $300 for the antibody. Sure, I could take the company’s word that it works in fluorescent staining techniques, but I need more proof. I need to see that it will work in my experiment and that means finding published images in the scientific literature.

Seeing, after all, is believing.

This process can take time.

For the antibody I’m looking for, there are at least three different versions (or clones).

I’ve spent a couple of hours trying to find just the right one, sifting through the “data.”

The antibody I’ve chosen, clone MEK14.7, seems to work in just about any experimental application. It’s rare to find an antibody that works in fresh samples as well as formalin-fixed samples, but this one does. Why? The molecule the antibody binds to in the tissue (called an antigen) often changes shape when fixed in formalin. The shape determines if the antibody can bind. If it changes, the antibody no longer recognizes (and binds) it.

Now, if I can find this antibody with the right dye on it, life will be good.

———————————————————————————————————————————————–

Update

If you look at the comments to this post, you will notice that Tom suggested the website 1DegreeBio. com  

I was unaware of this website, but it would have saved me some time in searching for an antibody.

I’m pretty sure I would try it first for any of my antibody searches.

The only thing lacking at this time (and it will improve over time as visitors add their two cents) is there were no ratings/reviews of the antibody reagents I looked at. This would be a big help, although I would caution that if the reviews are done by individual researchers, they should be taken with a grain of salt:  what works well in one researcher’s hands may not in the hands of another.

Why? It could be how the sample was handled before staining with the antibody (such as storage or fixation). It could be the incubation time of the antibody with the tissue — some labs do an overnight incubation in the cold, others (such as myself) incubate an hour or two at room temperature, and some incubate for 30 minutes at body temperature (37 degrees Celsius). These are only two examples, there are several steps in the staining process that could influence the antibody’s performance in an assay. 

Thanks for the suggestion, Tom! 🙂

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  1. Do you know about 1DegreeBio. They’re starting to rate & review antibodies.

    • I did NOT know about this website. Tried it out. No ratings or reviews, but it does have a nice database/search engine for antibodies/companies.

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