In splits

I have a small confession to make. When I said worked with, in the previous post, it’s not entirely true. I just stood by and watched while my guide separated them out into culture plates. I actually handled the cells on my own today. I diluted the cells and plated them all by myself. My guide was there to make sure I didn’t mess up, though.

Remember what I said about 293T needing TLC? It’s like this. As the cells grow, they multiply. And when they multiply it leads to crowding. See, 293T grows best when it has some surface to cling onto. But there’s only so much area on a single culture plate for them to grow on. Secondly, when crowding happens, there’s more waste products being released into the culture medium, and less nutrition available to each individual cells. That’s why we need to dilute the cells, to make sure there’s enough room for all of them.

Dilution isn’t a simple matter of just pouring more medium into the plate. It’s a cell culture, not orange squash. First, you have to remove the old medium, peel the cells off the plate (not literally) and then put a fraction of the cells you’ve recovered into a fresh plate with fresh medium. And all of this has to be carried out under sterile conditions. 293T is particularly sensitive to bacterial infection. (FYI, most culture media are banquets to bacteria and fungi. The media are so chock-full of nutrients it’s almost as if they beg to be contaminated)

The whole procedure took me around an hour and half, from start to finish. And I think I’ve done a reasonably good job. I won’t know till the 16th, though. That’s when the cells should have settled on the plate and started multiplying. Hope they’re OK. Fingers crossed!