Robust estimation of bacterial cell count from optical density

The iGEM 2018 interlab article is published today in Nature Communications Biology! This article, which I wrote about last October when we posted on bioRxiv, presents a cheap and easy protocol for estimating cell count and per-cell fluorescence on plate readers. This is so cheap (reagents <$1) and easy, as validated by hundreds of iGEM teams around the world, that I believe no paper should ever be accepted again if it has plate reader data in uncalibrated units, any more than we would accept a paper that measured length in cubits.

The core idea is pretty simple: basically, you calibrate against dilutions of silica microspheres with similar size and optical properties to cells. As long as your cells are in liquid culture that's not too opaque and the cells aren't doing anything really odd optically themselves, this should give a good relationship between optical density and cell count.

We validated this by combining our microsphere protocol and previously published fluorescence estimation protocol to get a close match between flow cytometer and plate reader estimates of per-cell fluorescence, which wouldn't work if either of the protocols was problematic.

 

Per-cell fluorescence from flow cytometer and plate reader (article Figure 5)

Ironically, the most challenging part of the whole publication was the author list. All the data was supplied by a lot of iGEMers: our consortium author list included approximately1400 people from about 250 teams all around the world.  I had to write scripts to manage the author list and to format and reformat it as we went back and forth with the journal to figure out how to match their formatting requirements. All said and done, however, I wouldn't have it any other way: I am proud to have been able to work with so many capable collaborators and with so many eager young contributors to synthetic biology.