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=Positive Control Dilution Test= The idea behind this is that the positive controls I have been using have been at levels of about 1 pmol, which would be approximately 10^11 molecules available for latch/padlock binding. This might be an issue, since the experiments I've been running have fewer available molecules. For example, 10,000 cells at 10,000 transcripts per cell (about as much as would ever exist) is still only 10^8 copies for detection, and that is assuming ALL detection probes (C probes or antibodies) have bound in pairs, which is unlikely. More reasonable copy numbers would be 10^7 (~thousand/cell), 10^6 (~hundred/cell) or even 10^5 (~ten/cell) for some. I will try to go as low as possible and see where the detection ends. {| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext> |- style="background-color:#B7DEE8;font-size:12pt;font-weight:bold" align="center" | width="65" height="27" | Tube No | width="83" | Dilution | width="68" | Molecules Available |- style="font-size:12pt" | align="center" height="15" valign="bottom" | 0 | align="center" align="center" valign="bottom" | 1 | align="center" align="center" valign="bottom" | 100000000000 |- style="background-color:#D9D9D9;font-size:12pt" | align="center" height="15" valign="bottom" | 1 | align="center" align="center" valign="bottom" | 10 | align="center" align="center" valign="bottom" | 10000000000 |- style="font-size:12pt" | align="center" height="15" valign="bottom" | 2 | align="center" align="center" valign="bottom" | 100 | align="center" align="center" valign="bottom" | 1.E+09 |- style="background-color:#D9D9D9;font-size:12pt" | align="center" height="15" valign="bottom" | 3 | align="center" align="center" valign="bottom" | 1000 | align="center" align="center" valign="bottom" | 1.E+08 |- style="font-size:12pt" | align="center" height="15" valign="bottom" | 4 | align="center" align="center" valign="bottom" | 10000 | align="center" align="center" valign="bottom" | 1.E+07 |- style="background-color:#D9D9D9;font-size:12pt" | align="center" height="15" valign="bottom" | 5 | align="center" align="center" valign="bottom" | 100000 | align="center" align="center" valign="bottom" | 1.E+06 |- style="font-size:12pt" | align="center" height="15" valign="bottom" | 6 | align="center" align="center" valign="bottom" | 1.E+06 | align="center" align="center" valign="bottom" | 1.E+05 |- style="background-color:#D9D9D9;font-size:12pt" | align="center" height="15" valign="bottom" | 7 | align="center" align="center" valign="bottom" | 1.E+07 | align="center" align="center" valign="bottom" | 10000 |- style="font-size:12pt" | align="center" height="15" valign="bottom" | 8 | align="center" align="center" valign="bottom" | 1.E+08 | align="center" align="center" valign="bottom" | 1000 |- style="background-color:#D9D9D9;font-size:12pt" | align="center" height="15" valign="bottom" | 9 | align="center" align="center" valign="bottom" | 1.E+09 | align="center" align="center" valign="bottom" | 100 |- style="font-size:12pt" | align="center" height="15" valign="bottom" | 10 | align="center" align="center" valign="bottom" | 1.E+10 | align="center" align="center" valign="bottom" | 10 |- style="background-color:#D9D9D9;font-size:12pt" | align="center" height="15" valign="bottom" | 11 | align="center" align="center" valign="bottom" | 1.E+11 | align="center" align="center" valign="bottom" | 1 |} I'll make the dilutions by diluting 5 uL of the previous dilution into 45 uL nfH2O. Hopefully this will be enough to prevent dilution artifacts until we get down near the lower limits. Since it is late in the afternoon already, I'll make the dilutions today and begin the experiment [[Daniel:Notebook/ComboLock/2016-8-27|tomorrow]]. [[Category:ComboLock]] [[Category:20160826]]
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