# Endy:Victor3 Calculating fluorescent protein synthesis

## Background subtraction

Subtract a media background, ${\displaystyle A_{media}}$, from the raw absorbance data, ${\displaystyle A_{raw}}$, and assume that the resulting data, ${\displaystyle A_{corrected}}$, is directly proportional to the number of cells in the well.

 ${\displaystyle {\frac {}{}}A_{corrected}=A_{raw}-A_{media}}$ ...Equation 1

Subtract a fluorescent protein-free cell background, ${\displaystyle G_{cells}}$, from the the raw fluorescent data, ${\displaystyle G_{raw}}$, and assume that the resulting data ${\displaystyle G_{corrected}}$ is proportional to the total number of GFP molecules in the well [immature GFP?].

 ${\displaystyle {\frac {}{}}G_{corrected}=G_{raw}-G_{cells}}$ ...Equation 2

## Unit conversion

Use standard calibration curves (see here for absorbance and here for fluorescence) to convert the background-corrected data into absolute units (CFU/well and GFP molecules per well). The calibration equations used are shown in Equations 3 & 4.

 ${\displaystyle {\frac {}{}}CFU=3.1e8*A_{corrected}-1.6e6}$ ...Equation 3 ${\displaystyle {\frac {}{}}GFP=7.0e8*G_{corrected}+6.0e11}$ ...Equation 4

## GFP synthesis rate calculations

To calculate the mean synthesis rate of GFP per cell, ${\displaystyle S_{cell}}$, assume the total GFP synthesis rate is equal to the time differential of ${\displaystyle GFP}$. ${\displaystyle S_{cell}}$ can be calculated as the total synthesis rate divided by ${\displaystyle CFU}$.

 ${\displaystyle {\frac {}{}}S_{total}={\frac {d[GFP]}{dt}}}$ ...Equation 5 ${\displaystyle {\frac {}{}}S_{cell}={\frac {S_{total}}{CFU}}}$ ...Equation 6