User:Madeleine Y. Bee/Notebook/Single Molecule Fluorescence/2013/06/03: Difference between revisions

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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span>
|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Measurements and Sample Prep for FCS</span>
|style="background-color: #F2F2F2" align="center"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
|style="background-color: #F2F2F2" align="center"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
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==Entry title==
==June 3, 2013==
* Insert content here...
===Absorbance and Fluorescence Data===
=====Finish preparing thiol DNA, AuNP and ThT sample=====
2[ThT]:1[DNA]→ 2.2245μM ThT<br.>
10% of 250μL=25μL (within 10% allows for accurate concentration comparison)<br.>
Total solution volume: 275μL<br.>
(2.2245×10<sup>-6</sup>M)(275×10<sup>-6</sup>L)=6.11738×10<sup>-10</sup>moles ThT<br.>
(6.11738×10<sup>-10</sup>moles)/(25×10<sup>-6</sup>L)=24.5μM ThT<br.>
<br.>
To make 24.5μM ThT:<br.>
(120μM)V=(24.5μM)(25μL)<br.>
V=5.1μL in 19.9μL PBS buffer<br.>
<br.>
Add 25μL of 24.5μM ThT to 250μL DNA/AuNP solution<br.>
New concentrations:
*thiol DNA- 1.011μM
*AuNP- 13.486μM
*ThT- 2.2245μM
<br.>
=====Data=====
[[Image:Abs_data_thiol_DNA,_AuNP,_MB_06032013.png]]
<br.>
<br.>
[[Image:Abs_data_thiol_DNA%2C_ThT_06032013.png]]
<br.>
<br.>
[[Image:Fluor_data_thiol_DNA,_AuNP,_MB_06032013.png]]
<br.>
<br.>
[[Image:Fluor_data_thiol_DNA,_AuNP,_ThT_06032013.png]]
 
====Notes====
Samples containing Molecular Beacon excited at 532nm<br.>
Samples containing ThT excited at 440nm<br.>
 
===Sample Preparation for FCS and Calculations===
=====Thiol DNA, MB, AuNP Solution Dilutions=====
*Starting Concentrations:
**AuNP- 9.03nM
**MB-1268nM
**Thiol DNA- 677.157nM
1. 1nM AuNPs<br.>
(9.03nM)V=(1nM)(30μL)<br.>
V=3.32μL in 26.68μL PBS buffer
*New Concentrations:
**AuNP- 1nM
**MB- 140.33nM
**Thiol DNA- 74.94nM
2. 500pM AuNPs<br.>
15μL 1nM solution in 15μL PBS buffer
*New Concentrations:
**AuNP- 500pM
**MB- 70.165nM
**Thiol DNA- 37.47nM
3. 100pM AuNPs<br.>
6μL 500pM solution in 24μL PBS buffer
*New Concentrations:
**AuNP- 100pM
**MB- 14.033nM
**Thiol DNA- 7.55nM
=====Thiol DNA, MB Solution Dilutions=====
*Starting Concentrations:
**MB- 1268nM
**Thiol DNA- 727.5nM
4. 140.33nM MB<br.>
3.32μL in 26.68μL PBS buffer
*New Concentrations:
**MB- 140.33nM
**80.51nM
5. 70.165nM MB<br.>
15μL 140.33nM MB in 15μL PBS buffer
*New Concentrations:
**MB- 70.165nM
**Thiol DNA- 40.26nM
6. 14.033nM MB<br.>
6μL 70.165nM MB in 24μL PBS buffer
*New Concentrations:
**MB- 14.033nM
**Thiol DNA- 40.26nM
=====MB Solution Dilutions=====
*Starting Concentrations:
**MB- 1268nM
7. 140.33nM MB<br.>
3.32μL in 26.68μL PBS buffer<br.>
<br.>
8. 70.165nM MB<br.>
15μL 140.33nM MB in 15μL PBS buffer<br.>
<br.>
9. 14.033nM MB<br.>
6μL 70.165nM MB in 24μL PBS buffer<br.>




Revision as of 07:10, 8 July 2013

Measurements and Sample Prep for FCS <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page
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June 3, 2013

Absorbance and Fluorescence Data

Finish preparing thiol DNA, AuNP and ThT sample

2[ThT]:1[DNA]→ 2.2245μM ThT<br.> 10% of 250μL=25μL (within 10% allows for accurate concentration comparison)<br.> Total solution volume: 275μL<br.> (2.2245×10-6M)(275×10-6L)=6.11738×10-10moles ThT<br.> (6.11738×10-10moles)/(25×10-6L)=24.5μM ThT<br.> <br.> To make 24.5μM ThT:<br.> (120μM)V=(24.5μM)(25μL)<br.> V=5.1μL in 19.9μL PBS buffer<br.> <br.> Add 25μL of 24.5μM ThT to 250μL DNA/AuNP solution<br.> New concentrations:

  • thiol DNA- 1.011μM
  • AuNP- 13.486μM
  • ThT- 2.2245μM

<br.>

Data

<br.> <br.> <br.> <br.> <br.> <br.>

Notes

Samples containing Molecular Beacon excited at 532nm<br.> Samples containing ThT excited at 440nm<br.>

Sample Preparation for FCS and Calculations

Thiol DNA, MB, AuNP Solution Dilutions
  • Starting Concentrations:
    • AuNP- 9.03nM
    • MB-1268nM
    • Thiol DNA- 677.157nM

1. 1nM AuNPs<br.> (9.03nM)V=(1nM)(30μL)<br.> V=3.32μL in 26.68μL PBS buffer

  • New Concentrations:
    • AuNP- 1nM
    • MB- 140.33nM
    • Thiol DNA- 74.94nM

2. 500pM AuNPs<br.> 15μL 1nM solution in 15μL PBS buffer

  • New Concentrations:
    • AuNP- 500pM
    • MB- 70.165nM
    • Thiol DNA- 37.47nM

3. 100pM AuNPs<br.> 6μL 500pM solution in 24μL PBS buffer

  • New Concentrations:
    • AuNP- 100pM
    • MB- 14.033nM
    • Thiol DNA- 7.55nM
Thiol DNA, MB Solution Dilutions
  • Starting Concentrations:
    • MB- 1268nM
    • Thiol DNA- 727.5nM

4. 140.33nM MB<br.> 3.32μL in 26.68μL PBS buffer

  • New Concentrations:
    • MB- 140.33nM
    • 80.51nM

5. 70.165nM MB<br.> 15μL 140.33nM MB in 15μL PBS buffer

  • New Concentrations:
    • MB- 70.165nM
    • Thiol DNA- 40.26nM

6. 14.033nM MB<br.> 6μL 70.165nM MB in 24μL PBS buffer

  • New Concentrations:
    • MB- 14.033nM
    • Thiol DNA- 40.26nM
MB Solution Dilutions
  • Starting Concentrations:
    • MB- 1268nM

7. 140.33nM MB<br.> 3.32μL in 26.68μL PBS buffer<br.> <br.> 8. 70.165nM MB<br.> 15μL 140.33nM MB in 15μL PBS buffer<br.> <br.> 9. 14.033nM MB<br.> 6μL 70.165nM MB in 24μL PBS buffer<br.>



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