Team:SJTU-BioX-Shanghai/Measurement
2019 iGEM teams are invited and encouraged to participate in the Measurement Study. Our team took part in this study which aimed to be one of the labs to make the calibration in different condition. In our case, we drew the Particle Standard Curve and the Fluorescence Standard Curve..
Fluorescence Standard Curve
Materials:
Fluorescein (provided in kit)
10ml 1xPBS pH 7.4-7.6 (phosphate buffered saline)
96 well plate black
Method:
1.Spin down fluorescein kit tube to make sure pellet is at the bottom of tube
2.Prepare 10x fluorescein stock solution (100μM) by resuspending fluorescein in 1mL of 1xPBS
3.Dilute the 10x fluorescein stock solution with 1xPBS to make a 1x fluorescein solution with concentration 10μM.
4.Prepare the serial dilutions of fluorescein:
Add 100μL of PBS into wells A2, B2, C2, D2....A12, B12, C12, D12
Add 200μL of fluorescein 1x stock solution into A1, B1, C1, D1
Transfer 100μL of fluorescein stock solution from A1 into A2
Mix A2 by pipetting up and down 3x and transfer 100μL into A3
Mix A3 by pipetting up and down 3x and transfer 100μL into A4
Mix A4 by pipetting up and down 3x and transfer 100μL into A5
Mix A5 by pipetting up and down 3x and transfer 100μL into A6
Mix A6 by pipetting up and down 3x and transfer 100μL into A7
Mix A7 by pipetting up and down 3x and transfer 100μL into A8
Mix A8 by pipetting up and down 3x and transfer 100μL into A9
Mix A9 by pipetting up and down 3x and transfer 100μL into A10
Mix A10 by pipetting up and down 3x and transfer 100μL into A11
Mix A11 by pipetting up and down 3x and transfer 100μL into liquid waste
Repeat dilution series for rows B, C, D
5.Measure fluorescence of all samples under microplate reader.
6.Import data into Excel and draw fluorescein standard curve.
Data Result:
Figure 1. Preliminary data on Fluorescence Standard Curve
Figure 2. Fluorescence Standard Curve
Particle Standard Curve
Materials:
300μL Silica beads - Microsphere suspension (provided in kit, 4.7 x 108 microspheres)
dd H2O
96 well plate
Method:
1.Obtain the tube labeled “Silica Beads” from the Interlab test kit and vortex vigorously for 30 seconds.
2.Immediately pipet 96μL microspheres into a 1.5 mL eppendof tube.
3.Add 904μL of dd H2O to the microspheres and Vortex well.
4.Prepare the serial dilutions of silica beads as before.
5.Measure OD600 of all samples under microplate reader.
6.Import data into Excel and draw standard curve.
4.Prepare the serial dilutions of silica beads as before.
Data Result:
Figure 3. Preliminary data on Particle Standard Curve
Figure 4. Particle Standard Curve
SJTU-BioX-Shanghai
Contact us: sjtuigem@gmail.com
Bio-X Institute, Shanghai Jiao Tong University, Dongchuan Rd. 800