> For the complete documentation index, see [llms.txt](https://help.sbtinstruments.com/llms.txt). Markdown versions of documentation pages are available by appending `.md` to page URLs; this page is available as [Markdown](https://help.sbtinstruments.com/inoc/why-use-bactobox-r.md). # Why use BactoBox® OD and McFarland drift with cell size. A plate count cannot show where your culture is. BactoBox® counts cells and shows the growth stage. Optical density (OD), McFarland standards and plate counts are the usual tools for standardizing inoculum. Each one leaves part of your dose unknown. ## OD and McFarland measure turbidity, not cells Both methods measure scattered light. Scattering depends on cell size as well as on concentration, and cells change size throughout a growth curve. So the factor that converts turbidity to colony forming units (CFU) changes with them.

Optical methods are strongly dependent on the size of objects. The two samples scatter the same amount of light, but the bacteria in exponential stage (left) are considerably bigger than the cells in stationary stage (right). Same OD but ~5× different cell concentrations.

We tracked one *E. coli* culture with BactoBox®, plate counts and OD600 at the same time points. The OD-to-CFU factor rose 6.6× across the run. Calibrate early in the curve and every later number reads too low. Calibrate late and every earlier number reads too high. The drift goes one way, so more replicates do not average it away. See [Drifts in OD to CFU ratios](/inoc/explainers/drifts-in-od-to-cfu-ratios.md). Cell size also differs between organisms and media. The common rule that McFarland 0.5 equals about 1.5 × 108 CFU/mL holds roughly for *E. coli*. Other organisms give other counts at the same turbidity, so each one needs its own calibration. ## A plate count cannot show where your culture is A plate count gives you the dose days after you used it. It also cannot tell you where the culture was on its growth curve. The same count can come from a culture that is still growing, or from one already in decline phase.
Growth curve of one culture, plate count against time. A dashed line joins two points with the same plate count, one at about 9 hours while the culture is still growing, one at about 23 hours in decline phase, where it is losing culturability. The active culture window, exponential to early stationary phase, is shaded as the window to dose from.

Illustration. One culture gives the same plate count twice. The first time it is still growing. The second time it is in decline phase and losing culturability. Dose from the shaded active culture.

In decline phase, cells lose culturability hour by hour. Harvest a few hours later than last time and you dose fewer viable cells. The plate count shows that you missed, but not why. An overnight culture is especially exposed, because it runs on the clock. A small change in seed, temperature or timing moves the harvest to the other side of the peak. ## Know the growth stage when you dose BactoBox® counts cells in about 2 minutes, with no calibration curve. In an active culture, from exponential to early stationary phase, effectively every cell it counts also forms a colony. See [Correlation with plate counts](/technical-documentation/performance/correlation-with-plate-counts.md).

Klebsiella aerogenes growth curve from shake flask with tryptic soy broth (TSB). BactoBox® cells (lavender) is practically superimposed on the plate counts (CFU, yellow).

Every BactoBox® measurement reports CIZE, the average cell size. In *E. coli*, CIZE peaked at 2.1 µm as exponential growth began, and fell below 1 µm when growth stopped. Map the growth curve of your strain once. After that, one measurement can also tell you whether a new culture is still active. See [Read the growth phase from cell size](/inoc/worked-examples/read-the-growth-phase-from-cell-size.md).

Illustration of cell size during a growth curve.

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