Klapperich Lab:Notebook/Lab Meeting Notes/2010/02/04: Difference between revisions

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=4 February 2010 Lab Meeting=
Qingqing Prospectus Practice Presentation.
'''‡ Announcements'''<br>
* Oakridge deadline is Confusing. The website says something different from the email. Have one from QQ- Anyone else? <br>
* Shichu Huang is here - WELCOME! <br>
<br>
'''‡ Flu R01:Integration''' <br>
* meetings every other M 1-3pm. 705.  <br>
† HDA (Lead: Jaephil, Team:Sonali)<br>
* Start planning R01 for Submission on '''6/5/10.''' MM, JD, CMK. <br>
* Jaephil trying out the lyophilizer with just water to save on HDA reagents. Then Sonali lyophilize HDA mixes +/- trehalose.(MM)<br>
*HDA at 65, 66, 67, 68, 69, 70C done with C diff ATCC DNA. Works at all except 70C. Less amplification as Temp increases (MM)<br>
*HDA negative water reactions all have primer dimers/non-specific band at 70bp for c diff HDA (MM)<br>
*Jaephil to make COP chips for LOD expt with C diff DNA. (MM) <br>
* 2nd HDA design - include RT, dry reagent, reduced evap loss, and parallel filling, etc <br>
† Sample Concentration (Lead: Jane, Team: Jaephil) <br>
* New design transfer to COP - ongoing
* Set up COMSOL and StarCD, look for governing equations for simulation of evaporation.  Ongoing <br>
† SPE Column Optimization for DNA/RNA. (Lead: Sonali, Team: Hussam, Jessie, Brendan) <br>
* Sonali put out a new RNA protocol (correct SPE batter, homemade buffers, less ethanol washing, RNA Secure) before break. <br>
* Protocol tested with 10ng, 1ng, 0.1ng Stratagene purified human RNA (MM) and 100 uL of 1:64 dilution ATCC MDCK sup Flu A/PR/8/34 (JC) <br>
* Human Total RNA see % yields at 50% and fraction 1 + fraction 2 = 4-5ng RNA out of 10ng with all changes. With high vol *ethanol wash but other protocol improvements, see 40% yield at all other concentrations. <br>
*For Flu virus see improved Ct value by 7 cycles in fraction 1 from multiple channels <br>
*Also detecting 1.4 logs more of virus at 1:64 JC virus prep than before. <br>
*Switching from lambda phage DNA to TaqMan RNase P detection [update] <br>
* Hussam Runs chip with RNA (confirm procedure is correct) [update]<br>
* Jessie will run chips with serial virus dilutions from 1:1 to 1:1,000,000 to re-do previous dilution experimnent with WHO SYBR green assay on chip.<br>
* Sonali will assay BIDMC virus sample with newest protocol with 0.7uM Silica and compare with Qiagen RNA extraction with CDC PCR assay <br>
† PCR - CMI  (Lead: Qingqing) <br>
* 6th Integrated chip with SPE+RT+PCR chip was designed. <br>
* "crack" issue is solved.
* It works for ATCC flu virus, however low efficiency.
* working on improving the efficiency
† PCR of C.Difficile DNA (Qingqing)<br>
* Meeting today at 3:30pm <br>
* Toxin A. <br>
* HDA dilution experiments for toxin A has been tested in tube.  The products has been tested in Bio-analyzer. 1pg is the limitation for the template, in order to get a detectable result.
* Toxin B.
*PCR assay works on chip with ATCC C.dif DNA.
* old SPE DNA has ct values above 30, on chip PCR with these DNA does not work either on 30 cycle chip nor on 40 cycle chip.
* Qiagen extracted DNA from Lisa on Jan 6 have ct values between 20-30.
* on chip PCR with lisa's DNA also does not work either on 30 cycle chip nor on 40 cycle chip.
   
* PCR1 Paper back to MCK this week. <br>
* CMK: PCR 1 draft. Analytical Chem. <br>
<br>
'''‡C. diff Project''' (Cathie, Sonali, Satish Singh, Lisa J., His post doc )<br>
* See email notes.
<br>
'''‡ Coulter Flu Fraunhofer Project (Lead: Sonali, Team: Sonali, Jessie, Cathie, CMI Folks, Qingqing)'''
* New meeting time, T, 9:30 am every other week. <br>
<br>
'''‡ Agilent Automated Sample Preparation (Lead: Alex)'''<br>
* Paper on HotDog in progress.  <br>
'''‡ COBRA (Lead: Jaephil, Team: Cathie, Jane for virus only, PHO folks)<br>'''
* New metal piece - First week of Dec.<br>
* Evap paper -CMK edited, back to JD and JZ this week <br>
<br>
'''‡ Biointerfaces group (Lead: MinCheol, Team: Cathie, MCK, Wong and Meller folks)'''<br>
'''* Cathie will submit paper inquiry.<br>'''
<br>
'''‡ CIMIT- Sepsis (Lead:Cathie, Team:TBA)'''<br>
* First samples collected. <br>
<br>
'''‡ PATH Grant (Lead:Cathie, Team:Frank, Sean, Mark, Jake Trueb (ME))''' <br>
* Submitted Gantt Document to PATH 11/25. <br>
* Frank: 1-week in-tube stabilization experiment successful. Extractions completed for 1-week storage.  PCR of eluted samples in progress (28 Jan).<br>
* Sean: Ready to start preparing the monoliths today (1.21) after the meeting.  Will be able to make it to the meeting by 3 every week.  Hussam will be training me on PCR next week.  Need bench training still <br>
* Mark: as of 26 Jan. has loaded 6 and 4 month trials with 10ng/straw.  Pending results of the 1ng/straw 0-day trial will load 2 month trial next week and shorter time periods in weeks to come.  Straw fabrication ongoing.<br>
<br>
'''‡ IIH Senior Project.'''<br>
* Ordering ELISA Kits
* Determining materials to test fluorescence
** Transparancy, COP, PDMS (gold-standard)
** material list from study (Xurography, Daniel A. Bartholomeusz, p 1367)
* Planned to reconnect with Jose, he is out of town until feb.
* Returning to the cutter plotter at MIT for 'origami testing'
** microscope?




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4 February 2010 Lab Meeting

Qingqing Prospectus Practice Presentation.

‡ Announcements

  • Oakridge deadline is Confusing. The website says something different from the email. Have one from QQ- Anyone else?
  • Shichu Huang is here - WELCOME!


‡ Flu R01:Integration

  • meetings every other M 1-3pm. 705.

† HDA (Lead: Jaephil, Team:Sonali)

  • Start planning R01 for Submission on 6/5/10. MM, JD, CMK.
  • Jaephil trying out the lyophilizer with just water to save on HDA reagents. Then Sonali lyophilize HDA mixes +/- trehalose.(MM)
  • HDA at 65, 66, 67, 68, 69, 70C done with C diff ATCC DNA. Works at all except 70C. Less amplification as Temp increases (MM)
  • HDA negative water reactions all have primer dimers/non-specific band at 70bp for c diff HDA (MM)
  • Jaephil to make COP chips for LOD expt with C diff DNA. (MM)
  • 2nd HDA design - include RT, dry reagent, reduced evap loss, and parallel filling, etc

† Sample Concentration (Lead: Jane, Team: Jaephil)

  • New design transfer to COP - ongoing
  • Set up COMSOL and StarCD, look for governing equations for simulation of evaporation. Ongoing

† SPE Column Optimization for DNA/RNA. (Lead: Sonali, Team: Hussam, Jessie, Brendan)

  • Sonali put out a new RNA protocol (correct SPE batter, homemade buffers, less ethanol washing, RNA Secure) before break.
  • Protocol tested with 10ng, 1ng, 0.1ng Stratagene purified human RNA (MM) and 100 uL of 1:64 dilution ATCC MDCK sup Flu A/PR/8/34 (JC)
  • Human Total RNA see % yields at 50% and fraction 1 + fraction 2 = 4-5ng RNA out of 10ng with all changes. With high vol *ethanol wash but other protocol improvements, see 40% yield at all other concentrations.
  • For Flu virus see improved Ct value by 7 cycles in fraction 1 from multiple channels
  • Also detecting 1.4 logs more of virus at 1:64 JC virus prep than before.
  • Switching from lambda phage DNA to TaqMan RNase P detection [update]
  • Hussam Runs chip with RNA (confirm procedure is correct) [update]
  • Jessie will run chips with serial virus dilutions from 1:1 to 1:1,000,000 to re-do previous dilution experimnent with WHO SYBR green assay on chip.
  • Sonali will assay BIDMC virus sample with newest protocol with 0.7uM Silica and compare with Qiagen RNA extraction with CDC PCR assay


† PCR - CMI (Lead: Qingqing)

  • 6th Integrated chip with SPE+RT+PCR chip was designed.
  • "crack" issue is solved.
  • It works for ATCC flu virus, however low efficiency.
  • working on improving the efficiency

† PCR of C.Difficile DNA (Qingqing)

  • Meeting today at 3:30pm
  • Toxin A.
  • HDA dilution experiments for toxin A has been tested in tube. The products has been tested in Bio-analyzer. 1pg is the limitation for the template, in order to get a detectable result.
  • Toxin B.
  • PCR assay works on chip with ATCC C.dif DNA.
  • old SPE DNA has ct values above 30, on chip PCR with these DNA does not work either on 30 cycle chip nor on 40 cycle chip.
  • Qiagen extracted DNA from Lisa on Jan 6 have ct values between 20-30.
  • on chip PCR with lisa's DNA also does not work either on 30 cycle chip nor on 40 cycle chip.
  • PCR1 Paper back to MCK this week.
  • CMK: PCR 1 draft. Analytical Chem.


‡C. diff Project (Cathie, Sonali, Satish Singh, Lisa J., His post doc )

  • See email notes.


‡ Coulter Flu Fraunhofer Project (Lead: Sonali, Team: Sonali, Jessie, Cathie, CMI Folks, Qingqing)

  • New meeting time, T, 9:30 am every other week.


‡ Agilent Automated Sample Preparation (Lead: Alex)

  • Paper on HotDog in progress.


‡ COBRA (Lead: Jaephil, Team: Cathie, Jane for virus only, PHO folks)

  • New metal piece - First week of Dec.
  • Evap paper -CMK edited, back to JD and JZ this week


‡ Biointerfaces group (Lead: MinCheol, Team: Cathie, MCK, Wong and Meller folks)
* Cathie will submit paper inquiry.
‡ CIMIT- Sepsis (Lead:Cathie, Team:TBA)

  • First samples collected.


‡ PATH Grant (Lead:Cathie, Team:Frank, Sean, Mark, Jake Trueb (ME))

  • Submitted Gantt Document to PATH 11/25.
  • Frank: 1-week in-tube stabilization experiment successful. Extractions completed for 1-week storage. PCR of eluted samples in progress (28 Jan).
  • Sean: Ready to start preparing the monoliths today (1.21) after the meeting. Will be able to make it to the meeting by 3 every week. Hussam will be training me on PCR next week. Need bench training still
  • Mark: as of 26 Jan. has loaded 6 and 4 month trials with 10ng/straw. Pending results of the 1ng/straw 0-day trial will load 2 month trial next week and shorter time periods in weeks to come. Straw fabrication ongoing.


‡ IIH Senior Project.

  • Ordering ELISA Kits
  • Determining materials to test fluorescence
    • Transparancy, COP, PDMS (gold-standard)
    • material list from study (Xurography, Daniel A. Bartholomeusz, p 1367)
  • Planned to reconnect with Jose, he is out of town until feb.
  • Returning to the cutter plotter at MIT for 'origami testing'
    • microscope?




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