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Revision as of 01:04, 16 October 2018

Template Title Template Title

Interlab Study
UMaryland's Interlab Study!

We did this year's iGEM InterLab study. It was tremendous.

We followed the protocol given by iGEM absolutely perfectly. Look at the beautiful protocol we followed here.

Download our excel file we actually used here.

Calibration

OD600 Reference Point - LUDOX Protocol

LUDOX CL-X H2O
Replicate 1 0.058 0.034
Replicate 2 0.056 0.035
Replicate 3 0.058 0.035
Replicate 4 0.058 0.035
Arithmetic Mean 0.058 0.035
Corrected Abs600 0.058
Reference OD600 0.058
OD600/Abs600 0.058

Particle Standard Curve - Microsphere Protocol

Number of Particles 2.35×108 1.18×108 5.88×107 2.94×107 1.47×107 7.35×106 3.68×106 1.84×106 9.19×105 4.60×105 2.30×105 0
Replicate 1 0.716 0.322 0.191 0.095 0.067 0.059 0.039 0.036 0.036 0.035 0.034 0.036
Replicate 2 0.406 0.359 0.212 0.111 0.072 0.051 0.044 0.036 0.036 0.035 0.034 0.034
Replicate 3 0.587 0.347 0.203 0.116 0.075 - 0.041 0.038 0.038 0.036 0.035 0.034
Replicate 4 0.815 0.419 0.219 0.123 0.081 0.051 0.048 0.040 0.038 0.037 0.036 0.035
Arithmetic Mean 0.631 0.364 0.206 0.111 0.074 0.054 0.043 0.038 0.037 0.036 0.035 0.035
Arithmetic Standard Deviation 0.177 0.038 0.012 0.012 0.006 0.005 0.004 0.002 0.001 0.001 0.001 0.001
Arithmetic Net Mean 0.596 0.329 0.172 0.077 0.039 0.019 0.008 0.003 0.002 0.001 0.000
interlab particle standard curve interlab particle standard curve
Number of Particles 2.35×108 1.18×108 5.88×107 2.94×107 1.47×107 7.35×106 3.68×106 1.84×106 9.19×105 4.60×105 2.30×105
µM Fluorescein/a.u. 3.95×108 3.57×108 3.43×108 3.84×108 3.77×108 3.89×108 4.46×108 6.68×108 4.80×108 4.60×108 -
MEFL/a.u. 3.70×108

Fluorescence Standard Curve - Fluorescein Protocol

Fluorescein µM 10 5 2.5 1.25 0.625 0.313 0.156 0.078 0.039 0.0195 0.0098 0
Replicate 1 27845 21611 15480 10321 5979 3376 1806 891 442 244 122 2
Replicate 2 27711 19390 16110 10742 6223 3240 1756 895 467 227 115 2
Replicate 3 27686 21709 15590 9458 5304 2972 1350 711 481 101 160 3
Replicate 4 27431 21442 15379 9474 5366 2703 1427 716 359 197 115 3
Arithmetic Mean 27668 21038 15640 10000 5718 3073 1585 803 437 192 128 2.5
Arithmetic Standard Deviation 172.8 1104 325.1 638.8 454.0 298.2 229.7 103.7 54.60 63.86 21.59 0.5774
Arithmetic Net Mean 27666 21036 15637 9996 5716 3070 1582 801 435 190 126
interlab particle standard curve interlab particle standard curve
Flourescein µM 10.00 5.00 2.50 1.25 0.63 0.31 0.16 0.08 0.04 0.02 0.01
µM Fluorescein/a.u. 3.61×10-4 2.38×10-4 1.60×10-4 1.25×10-4 1.09×10-4 1.02×10-4 9.88×10-5 9.76×10-5 8.99×10-5 1.03×10-4 7.78×10-5
Mean µM Fluorescein/a.u. 1.47×10-4
MEFL/a.u. 8.84×108

Cell Measurement

Plate Reader Raw Measurement

Fluorescence Raw Readings

Hour 0:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6 LB + Chloro. (blank)
Colony 1 Replicate 1 23 43 63 44 24 84 63 55 23
Colony 1 Replicate 2 25 36 51 36 21 69 52 44 20
Colony 1 Replicate 3 25 35 51 38 23 70 52 44 20
Colony 1 Replicate 4 19 37 54 35 23 70 53 48 21
Colony 2 Replicate 1 25 35 80 29 25 74 68 38 21
Colony 2 Replicate 2 28 34 81 31 25 75 69 39 22
Colony 2 Replicate 3 27 35 83 32 25 75 68 40 21
Colony 2 Replicate 4 31 46 105 40 34 101 90 52 27
Hour 6:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6 LB + Chloro. (blank)
Colony 1 Replicate 1 31 288 447 251 37 132 111 205 24
Colony 1 Replicate 2 36 266 382 213 36 112 93 186 20
Colony 1 Replicate 3 35 252 382 216 37 108 102 178 21
Colony 1 Replicate 4 36 260 381 216 39 113 93 179 22
Colony 2 Replicate 1 36 260 381 216 39 116 140 161 24
Colony 2 Replicate 2 39 215 340 152 110 121 137 163 24
Colony 2 Replicate 3 37 210 348 150 108 117 139 161 23
Colony 2 Replicate 4 41 258 415 175 130 150 183 198 23

Abs600 Raw Readings

Hour 0:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6 LB + Chloro. (blank)
Colony 1 Replicate 1 0.077 0.073 0.074 0.078 0.075 0.068 0.068 0.068 0.060
Colony 1 Replicate 2 0.079 0.075 0.069 0.077 0.075 0.074 0.076 0.074 0.054
Colony 1 Replicate 3 0.077 0.074 0.062 0.078 0.076 0.077 0.077 0.077 0.059
Colony 1 Replicate 4 0.068 0.076 0.076 0.080 0.074 0.071 0.073 0.077 0.057
Colony 2 Replicate 1 0.081 0.077 0.084 0.077 0.072 0.074 0.079 0.078 0.060
Colony 2 Replicate 2 0.085 0.075 0.083 0.079 0.072 0.074 0.076 0.082 0.059
Colony 2 Replicate 3 0.081 0.076 0.083 0.081 0.074 0.074 0.088 0.081 0.057
Colony 2 Replicate 4 0.086 0.075 0.083 0.075 0.072 0.074 0.072 0.082 0.056
Hour 6:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6 LB + Chloro. (blank)
Colony 1 Replicate 1 0.445 0.437 0.258 0.447 0.487 0.108 0.329 0.459 0.062
Colony 1 Replicate 2 0.456 0.475 0.261 0.444 0.518 0.140 0.324 0.477 0.059
Colony 1 Replicate 3 0.473 0.433 0.270 0.450 0.524 0.109 0.353 0.450 0.063
Colony 1 Replicate 4 0.475 0.431 0.254 0.444 0.535 0.109 0.328 0.438 0.059
Colony 2 Replicate 1 0.456 0.414 0.229 0.459 0.444 0.115 0.112 0.463 0.060
Colony 2 Replicate 2 0.411 0.429 0.243 0.452 0.460 0.119 0.106 0.453 0.058
Colony 2 Replicate 3 0.435 0.418 0.234 0.474 0.460 0.120 0.113 0.455 0.063
Colony 2 Replicate 4 0.425 0.424 0.248 0.470 0.450 0.121 0.115 0.466 0.061

Fluoresence per OD

µM Flourescin/OD

Hour 0:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6
Colony 1, Replicate 1 0.000 0.078 0.145 0.059 0.003 0.386 0.253 0.203
Colony 1, Replicate 2 0.010 0.039 0.105 0.035 0.002 0.124 0.074 0.061
Colony 1, Replicate 3 0.014 0.039 0.105 0.035 0.002 0.124 0.074 0.061
Colony 1, Replicate 4 0.014 0.051 0.121 0.048 0.009 0.141 0.090 0.068
Colony 2, Replicate 1 -0.009 0.043 0.088 0.031 0.006 0.177 0.101 0.068
Colony 2, Replicate 2 0.010 0.042 0.125 0.023 0.012 0.179 0.140 0.037
Colony 2, Replicate 3 0.013 0.037 0.121 0.023 0.012 0.161 0.077 0.040
Colony 2, Replicate 4 0.007 0.051 0.146 0.035 0.022 0.208 0.199 0.049
Hour 6:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6
Colony 1, Replicate 1 0.001 0.036 0.109 0.030 0.002 0.119 0.017 0.023
Colony 1, Replicate 2 0.002 0.030 0.091 0.025 0.002 0.058 0.014 0.020
Colony 1, Replicate 3 0.002 0.032 0.089 0.026 0.002 0.092 0.013 0.021
Colony 1, Replicate 4 0.002 0.032 0.089 0.026 0.002 0.092 0.013 0.021
Colony 2, Replicate 1 0.002 0.026 0.091 0.015 0.011 0.085 0.113 0.017
Colony 2, Replicate 2 0.002 0.026 0.087 0.016 0.011 0.081 0.119 0.018
Colony 2, Replicate 3 0.002 0.027 0.096 0.016 0.011 0.084 0.118 0.018
Colony 2, Replicate 4 0.003 0.033 0.106 0.019 0.014 0.107 0.150 0.022

Net Flourescin a.u.

Hour 0:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6
Colony 1, Replicate 1 0 20 40 21 1 61 40 32
Colony 1, Replicate 2 5 16 31 16 1 49 32 24
Colony 1, Replicate 3 5 15 31 18 3 50 32 24
Colony 1, Replicate 4 -2 16 33 14 2 49 32 27
Colony 2, Replicate 1 4 14 59 8 4 53 47 17
Colony 2, Replicate 2 6 12 59 9 3 53 47 17
Colony 2, Replicate 3 6 14 62 11 4 54 47 19
Colony 2, Replicate 4 4 14 78 13 7 74 63 25
Hour 6:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6
Colony 1, Replicate 1 7 264 423 227 13 108 87 181
Colony 1, Replicate 2 16 261 263 193 16 92 73 166
Colony 1, Replicate 3 14 231 362 195 16 87 81 157
Colony 1, Replicate 4 14 238 359 194 17 91 71 157
Colony 2, Replicate 1 12 184 305 121 81 92 116 137
Colony 2, Replicate 2 15 191 316 128 86 97 113 139
Colony 2, Replicate 3 14 187 325 127 85 94 116 138
Colony 2, Replicate 4 18 235 392 152 107 127 160 175
Hour 0:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6
Colony 1, Replicate 1 0.017 0.013 0.014 0.018 0.015 0.008 0.008 0.008
Colony 1, Replicate 2 0.025 0.021 0.015 0.023 0.021 0.020 0.022 0.020
Colony 1, Replicate 3 0.018 0.015 0.013 0.019 0.017 0.018 0.018 0.018
Colony 1, Replicate 4 0.011 0.019 0.019 0.023 0.017 0.014 0.016 0.020
Colony 2, Replicate 1 0.021 0.017 0.024 0.017 0.012 0.014 0.019 0.018
Colony 2, Replicate 2 0.026 0.016 0.024 0.020 0.013 0.015 0.017 0.023
Colony 2, Replicate 3 0.024 0.019 0.026 0.024 0.017 0.017 0.031 0.024
Colony 2, Replicate 4 0.030 0.019 0.027 0.019 0.016 0.018 0.016 0.026
Hour 6:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6
Colony 1, Replicate 1 0.383 0.375 0.196 0.385 0.425 0.046 0.267 0.397
Colony 1, Replicate 2 0.397 0.416 0.202 0.385 0.459 0.081 0.265 0.418
Colony 1, Replicate 3 0.410 0.370 0.207 0.387 0.461 0.046 0.290 0.387
Colony 1, Replicate 4 0.416 0.372 0.195 0.385 0.476 0.050 0.269 0.379
Colony 2, Replicate 1 0.396 0.354 0.169 0.399 0.384 0.055 0.052 0.403
Colony 2, Replicate 2 0.353 0.371 0.185 0.394 0.402 0.061 0.048 0.395
Colony 2, Replicate 3 0.372 0.355 0.171 0.411 0.397 0.057 0.050 0.392
Colony 2, Replicate 4 0.364 0.363 0.187 0.409 0.389 0.060 0.054 0.405

Fluorescence per Particle

MEFL/particle

Hour 0:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6
Colony 1, Replicate 1 0 3.67×103 6.82×103 2.79×103 1.59×102 1.82×104 1.19×104 9.55×103
Colony 1, Replicate 2 4.78×102 1.82×103 4.93×103 1.66×103 1.14×102 5.85×103 3.47×103 2.87×103
Colony 1, Replicate 3 6.63×102 2.39×103 5.69×103 2.26×103 4.21×102 6.63×103 4.24×103 3.18×103
Colony 1, Replicate 4 -4.34×102 2.01×103 4.15×103 1.45×103 2.81×102 8.36×103 4.78×103 3.22×103
Colony 2, Replicate 1 4.55×102 1.97×103 5.87×103 1.12×103 7.96×102 9.04×103 5.91×103 2.26×103
Colony 2, Replicate 2 5.51×102 1.79×103 5.87×103 1.07×103 5.51×102 8.44×103 6.60×103 1.76×103
Colony 2, Replicate 3 5.97×102 1.76×103 5.69×103 1.09×103 5.62×102 7.58×103 3.62×103 1.89×103
Colony 2, Replicate 4 3.18×102 2.39×103 6.90×103 1.63×103 1.04×103 9.82×103 9.40×103 2.03×103
Hour 6:
Negative Control Positive Control Device 1 Device 2 Device 3 Device 4 Device 5 Device 6
Colony 1, Replicate 1 4.36×101 1.68×103 5.15×103 1.41×103 7.30×101 5.61×103 7.78×102 1.09×103
Colony 1, Replicate 2 9.62×101 1.41×103 2.28×103 1.20×103 8.32×101 2.71×103 6.58×102 9.48×102
Colony 1, Replicate 3 8.15×101 1.49×103 4.18×103 1.20×103 8.29×101 4.52×103 6.67×102 9.69×102
Colony 1, Replicate 4 8.04×101 1.53×103 4.40×103 1.20×103 8.53×102 4.35×103 6.30×102 9.89×103
Colony 2, Replicate 1 7.24×101 1.24×103 4.31×103 7.24×102 5.04×102 3.99×103 5.33×103 8.12×102
Colony 2, Replicate 2 1.01×102 1.23×103 4.08×103 7.76×102 5.11×102 3.80×103 5.62×103 8.40×102
Colony 2, Replicate 3 8.99×101 1.26×103 4.54×103 7.38×102 5.11×102 3.94×103 5.54×103 8.41×102
Colony 2, Replicate 4 1.18×102 1.55×103 5.01×103 8.87×102 6.57×102 5.05×103 7.07×103 1.03×103

Colony Forming Units per 0.1 OD600 E. coli Cultures

Colonies per Plate

Positive Control

Colony 1, Replicate 1 Colony 1, Replicate 2 Colony 1, Replicate 3 Colony 2, Replicate 1 Colony 2, Replicate 2 Colony 2, Replicate 3
Dilution 3 >300 >300 >300 >300 >300 >300
Dilution 4 64 129 127 9 143 80
Dilution 5 13 8 12 5 18 5

Negative Control

Colony 1, Replicate 1 Colony 1, Replicate 2 Colony 1, Replicate 3 Colony 2, Replicate 1 Colony 2, Replicate 2 Colony 2, Replicate 3
Dilution 3 >300 >300 >300 157 >300 >300
Dilution 4 >300 >300 92 259 5 3
Dilution 5 41 33 14 11 0 0

Colony Forming Units per mL

Positive Control

Colony 1, Replicate 1 Colony 1, Replicate 2 Colony 1, Replicate 3 Colony 2, Replicate 1 Colony 2, Replicate 2 Colony 2, Replicate 3
Dilution 3 - - - - - -
Dilution 4 5.12×107 1.03×108 1.02×108 7.20×106 1.14×108 6.40×107
Dilution 5 1.04×108 6.40×107 9.60×107 4.00×107 1.44×108 4.00×107

Negative Control

Colony 1, Replicate 1 Colony 1, Replicate 2 Colony 1, Replicate 3 Colony 2, Replicate 1 Colony 2, Replicate 2 Colony 2, Replicate 3
Dilution 3 - - - 1.26×107 - -
Dilution 4 - - 7.36×107 2.07×108 4.00×106 2.40×106
Dilution 5 3.28×108 2.64×108 1.12×108 8.80×107 0 0

Miscellaneous Information

The 96 well plate we used was clear. Some of the flourescence data might be affected due to bleeding of flourescence from certain wells to another.

yeah we realize this is not so pretty. we're working on it.

contact us at umarylandigem@gmail.com

Contact Us
umarylandigem@gmail.com
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4094 Campus Dr, College Park, MD 20742

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