User:Karlena L. Brown/Notebook/PVOH Research/2013/02/13: Difference between revisions

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==PVOH Film Preparations==
==OBJECTIVES==
'''Prepared PVOH films in water:'''
* Run fluorescence analysis on all hydrogel samples prepared with Rhodamine 6G dye (dye leaching out of medium anaylsis)
* In 10mL beaker, weigh out ~ 0.5 grams PVOH (MW 130,000)
* Grind microsphere PVOH 146K 110% CEC Laponite w/ DMHXLBR into a smaller sample
'''(Actual Mass = 0.4934g)'''
* Re-evaluate strategy as to how to prepare more microspheres more efficiently
* Then, using a graduated cylinder add ~ 3 mL H<sub>2</sub>O to the beaker
* In another 10mL beaker, weigh out ~ 1.0 gram PVOH (MW 130,000)
'''(Actual Mass = 0.9965g)'''
* Then, using a graduated cylinder add ~ 5 mL H<sub>2</sub>O to the beaker


'''Standard PVOH Film Protocol:'''
==Fluorescence Analysis Instructions==
* After adding and combining PVOH (MW 130,000) in small beakers with H<sub>2</sub>O, add stir bars and prepare to stir solution.
# Start the FLWin Lab program
* On hot plate, stir and heat both beaker solutions at 70-80°C for ~ 12 min or until PVOH dissolves.
# Turn on the machine by clicking the on button on the side of the instrument
* Once PVOH solids thoroughly dissolve in solution, pour each solution in a Teflon dish to sit, cool, and dry in a fume hood for ~ 1 day.
# Go to and select the Graph Button Command
# Afterwards select and the Setup Parameters Tab
# Within the Setup Parameters Tab, select the Emission Tab
# Set up parameters for fluorescence analysis by determining emission and excitation ranges
# Afterwards, turn on the lamp source by clicking on the Lamp Button in the instrument window
# Pour ~ 1-2mL of sample into a glass cuvette marked for fluorescence
# Then place the sample filled cuvette within the sample holder
# Next, in the Graph Window select the Lamp Button Icon to run the sample
# Once the sample has finished running, back in the main window select the Save As Command
# Finally officially save the sample measurement and file information as a text file under '''ASC II'''
# Create folder a to save the information under '''MK'''


'''Notes:'''
===Fluorescence Limit of Detection Calculations===
* An additional 2-3 mL H<sub>2</sub>O was added to each solution to aid the dissolving of the PVOH
'''Fluorescent Standards For Limit of Detection: 0.25μM, 0.5μM, 1μM, 92μM, & 165μM'''
* Heat was reduced when solutions began to boil too rapidly
* While transferring solutions to Teflon dishes, loss of sample within the small beakers (sample loss = incomplete sample transfer)


'''0.25μM Rhodamine 6G Dye Concentration'''
  M<sub>1</sub>V<sub>1</sub> = M<sub>2</sub>V<sub>2</sub>
  0.25μM (RG6)x 5mL = (0.5μM)V<sub>2</sub>    V<sub>2</sub> = 2.5mL
'''0.5μM Rhodamine 6G Dye Concentration'''
  M<sub>1</sub>V<sub>1</sub> = M<sub>2</sub>V<sub>2</sub>
  0.5μM (RG6)x 10mL = (92μM)V<sub>2</sub>    V<sub>2</sub> = 54.34μL
'''1μM Rhodamine 6G Dye Concentration'''
  M<sub>1</sub>V<sub>1</sub> = M<sub>2</sub>V<sub>2</sub>
  1μM (RG6)x 25mL = (92μM)V<sub>2</sub>    V<sub>2</sub> = 271.74μL
==Notes==
* Microsphere sample PVOH 146K 110% CEC Laponite w/ DMHXLBR actually came out as a hunk of material rather than microspheres
* For fluorescent analysis, several Rhodamine 6G dye standards were prepared in order to determine the limit of detection
* Every sample previously prepared on 1/30/13 were analyzed because Rhodamine 6G dye was not crosslinked into hydrogel material
* Specific parameters used to analyze fluorescence of hydrogel samples containing Rhodamine 6G dye included the following:
    # Excitation: 480nm
    # Emission Range: 500-650nm
    # Excitation Slit Width: 10
    # Emission Slit Width: 10
    # Scan Speed: 1200
   


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Revision as of 22:58, 17 February 2013

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OBJECTIVES

  • Run fluorescence analysis on all hydrogel samples prepared with Rhodamine 6G dye (dye leaching out of medium anaylsis)
  • Grind microsphere PVOH 146K 110% CEC Laponite w/ DMHXLBR into a smaller sample
  • Re-evaluate strategy as to how to prepare more microspheres more efficiently

Fluorescence Analysis Instructions

  1. Start the FLWin Lab program
  2. Turn on the machine by clicking the on button on the side of the instrument
  3. Go to and select the Graph Button Command
  4. Afterwards select and the Setup Parameters Tab
  5. Within the Setup Parameters Tab, select the Emission Tab
  6. Set up parameters for fluorescence analysis by determining emission and excitation ranges
  7. Afterwards, turn on the lamp source by clicking on the Lamp Button in the instrument window
  8. Pour ~ 1-2mL of sample into a glass cuvette marked for fluorescence
  9. Then place the sample filled cuvette within the sample holder
  10. Next, in the Graph Window select the Lamp Button Icon to run the sample
  11. Once the sample has finished running, back in the main window select the Save As Command
  12. Finally officially save the sample measurement and file information as a text file under ASC II
  13. Create folder a to save the information under MK

Fluorescence Limit of Detection Calculations

Fluorescent Standards For Limit of Detection: 0.25μM, 0.5μM, 1μM, 92μM, & 165μM

0.25μM Rhodamine 6G Dye Concentration 

  M1V1 = M2V2
  0.25μM (RG6)x 5mL = (0.5μM)V2    V2 = 2.5mL

0.5μM Rhodamine 6G Dye Concentration 

  M1V1 = M2V2
  0.5μM (RG6)x 10mL = (92μM)V2    V2 = 54.34μL

1μM Rhodamine 6G Dye Concentration 

  M1V1 = M2V2
  1μM (RG6)x 25mL = (92μM)V2    V2 = 271.74μL

Notes

  • Microsphere sample PVOH 146K 110% CEC Laponite w/ DMHXLBR actually came out as a hunk of material rather than microspheres
  • For fluorescent analysis, several Rhodamine 6G dye standards were prepared in order to determine the limit of detection
  • Every sample previously prepared on 1/30/13 were analyzed because Rhodamine 6G dye was not crosslinked into hydrogel material
  • Specific parameters used to analyze fluorescence of hydrogel samples containing Rhodamine 6G dye included the following:
    # Excitation: 480nm
    # Emission Range: 500-650nm
    # Excitation Slit Width: 10
    # Emission Slit Width: 10
    # Scan Speed: 1200


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