Measurement of optical density of microbes by multi‐light path transmission method

Optical density (OD) is an important indicator of microbial density, and a commonly used variable in growth curves to express the growth of microbial culture. However, OD values show a linear relationship with bacterial concentration only at low concentrations. When the cell density is high, the relationship loses linearity, and serial dilution is needed to obtain readings of better accuracy. Here, we show that measuring OD values using shorter light paths is in close equivalence to measuring OD values of the cell culture with corresponding dilution. By measuring three different light paths simultaneously, accurate OD values can be easily obtained from low to high cell density. Using this method, growth curves of <i>Escherichia coli</i>, <i>Staphylococcus aureus</i>, and <i>Pichia pastoris</i> are measured with higher accuracy. To further simplify the process, an l-shaped cuvette and a corresponding turbidimeter are designed specifically for OD value measurement based on the multi-light path transmission method.