Team:Warwick/InterLab

Ripple

InterLab

Goals

To establish a common conversion factor from fluorescence/OD600 to CFU/ml, which acts as a more absolute unit than the relative unit of OD600. We were supplied with six plasmid test constructs each containing an RBS to express a composite part, as well as positive and negative controls. We also received a Standard Measurement Kit in order to create standard curves for converting Abs600 to OD600, an absorption standard curve and a fluorescence standard curve.

Device Part Number
Negative control BBa_R0040
Positive Control BBa_I20270
Test Device 1 BBa_J364000
Test Device 2 BBa_J364001
Test Device 3 BBa_J364002
Test Device 4 BBa_J364007
Test Device 5 BBa_J364008
Test Device 6 BBa_J364009

Results

Volumes of culture and media required for 0.02 starting OD600
Target Abs600 = 0.02, Target volume (mL) = 6

Sample Abs600 Volume of culture(mL) Volume of LB + Cam media (mL)
Positive control colony 1 0.1247 2.26 3.74
Positive control colony 2 0.12595 2.21 3.79
Negative control colony 1 0.13665 1.85 4.15
Negative control colony 2 0.13305 1.96 4.04
Device 1 colony 1 0.0959 1.96 4.04
Device 1 colony 2 0.11865 2.56 3.44
Device 2 colony 1 0.161 1.34 4.66
Device 2 colony 2 0.14855 1.56 4.44
Device 3 colony 1 0.1269 2.17 3.83
Device 3 colony 2 0.14105 1.73 4.27
Device 4 colony 1 0.11275 2.92 3.08
Device 4 colony 2 0.0959 1.96 4.04
Device 5 colony 1 0.1061 3.49 2.51
Device 5 colony 2 0.11295 2.91 3.09
Device 6 colony 1 0.1378 1.82 4.18
Device 6 colony 2 0.11935 2.52 3.48

Volumes of culture and media required for 0.1 OD600

Target Abs600 = 0.1, Target volume = 1000μl


Sample Abs600 Volume of culture (μl) Volume of LB + Cam media (μl) Average Actual Abs600
Positive control colony 1 0.115 292 708 0.0829
Positive control colony 2 0.1126 309 691 0.0645
Negative control colony 1 0.1312 212 788 0.0584
Negative control colony 2 0.1287 221 779 0.0571

OD600

LUDOX CL-X H2O
Replicate 1 0.090 0.066
Replicate 2 0.082 0.071
Replicate 3 0.088 0.070
Replicate 4 0.086 0.068
Arith Mean 0.087 0.069
Corrected Abs600 0.018 -
Reference OD600 0.063 -
OD600/Abs600 3.554 -

Microsphere Standard Curve

Fluorescein Standard Curve

Raw Plate Readings

Fluorescence/OD600

Fluorescence/Particle

CFU/ml

Colonies 8*108 dilution CFU/ml
Positive control colony 1.1 327 2.616E+11
Positive control colony 1.2 258 2.064E+11
Positive control colony 1.3 368 2.944E+11
Positive control colony 2.1 329 2.632E+11
Positive control colony 2.2 231 1.848E+11
Positive control colony 2.3 341 2.728E+11
Negative control colony 1.1 284 2.272E+11
Negative control colony 1.2 298 2.384E+11
Negative control colony 1.3 248 1.984E+11
Negative control colony 2.1 274 2.192E+11
Negative control colony 2.2 344 2.752E+11
Negative control colony 2.3 363 2.904E+11