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As a PhD researcher working on biological imaging agents, live cell imaging can be a right pain.
Despite the joys of antiquated equipment, system failures, or using the wrong laser, the actual aspect of cell imaging takes time and effort.
There’s something quite miraculous and wonderful watching a 20 µM
bundle of proteins, lipids and carbohydrates (with a large amount of water added) growing and absorbing nutrients.
Add in some imaging agents, or dyes, and the cells glow a variety of colours that allow you to begin to unravel the inner workings of the cell.
However, getting to this step can be drawn out, full of swearing, and sometimes tears. I’m sure there are a multitude of reasons as to why imaging cells sucks, but here are the 6 that I can think of.
“The alarm for the CO2 incubator went off on the weekend” is normally the dreaded phrase any scientist working with cells fears.
After all your careful planning: ensuring the hood was kept sterile; buying in new reagents; carefully culturing your cells for the past couple of days – all for nothing as you go to check the incubator and find your cells unattached and floating (unhappily) in your media… dead.
This has happened more frequently than I’d like to mention, though other issues with cell incubators can be: the temperature dropped too much; there wasn’t any water to generate humidity; someone left the incubator open and had muted the alarm…
Sterile labs and good lab practice are essential to culturing cells for imaging. Keeping reagents un-contaminated in a busy cell lab is both an art, and a chore.
It only takes one swipe of the pipette tip touching an area that you failed to rigorously clean and you might have a nice little bacteria colony growing in your cells. Nothing like looking under the microscope and little rods and dots zipping around in your “sterile” media containing cells.
Overall, this is a very infrequent occurrence that I’ve not had to deal with for a couple of years now.
Most labs I’ve worked in have always had a stock of Hela cells on the go. Easy to grow, resistant to a lot of things, these adherent immortal cells have found a place in history for several reasons (both good and bad).
Yet, they aren’t always representative of what your compounds are doing. Therefore, you normally need a second set of cells as a control – which come with their own growing conditions (and stresses).
Or perhaps you needed adherent cells vs suspension cells, and therefore your experiments fail because the uptake process isn’t the same.
You go to grab your 8 well slide that’s been incubating cells with your imaging agent for 24 hours. You’ve set everything up ready to image, and you go to check what the cells look like.
Everything is dead. You check to make sure that nothing has happened with the incubator, or that you’ve had a contamination. Nope, nothing. Finally, you check the control well and everything is… fine.
So that amazing compound you’ve synthesized is just a toxic mess. Even low concentrations cause your cells to undergo apoptosis, and it can occur within hours.
So back to the drawing board with that experiment!
So, you’ve gotten as far as the microscope. Cells are fine, your compounds aren’t killing the cells.
The imaging software is working, if slow. Everything is set, and you are ready to image. Hours go by, and what you expect to see isn’t happening.
Your bright red compounds aren’t fluorescing correctly. The Hoechst stain of the nucleus also doesn’t seem to be working correctly, and not very bright.
You spend another hour trying to work out why your experiment hasn’t worked, then you realize it’s the lasers.
You’ve got the wrong laser and it’s not exciting your samples correctly!
You’ve managed to get this far, but now you want to have everything fluorescently labelled. You’ve fixed your cells so that you can maintain them as they were.
You want to know where your compounds lie in relation to the nucleus. You’ve decided to do a cytoskeleton stain as well. However, you've forgotten to check where each of the stains absorbs and emits.
Is that green the cytoskeleton stain, or is it your compound?
Why is the nucleus stain leeching?
Why is that green as well?
Why is the Z-stack program not working?
Why does live cell imaging have to be so difficult?!
At Incuvers, our CEO Sebastian suffered the exact struggles that Struan did, spending years of his PhD research capturing just a few time lapse videos of his cells.
That's why Incuvers has developed a solution, IRIS.
What if your incubator did live cell imaging for you? What if you could check on your cells from your phone, instead of risking removing them from the incubator?