Notes on calculations of concentrations from gel images 4-11-13

2015-01-13

azim58 - Notes on calculations of concentrations from gel images 4-11-13

Total Intensity of band 3rd from bottom (300 bp)

40591
10 uL ladder was loaded so this band corresponds to 16.5 ng

Conversion factor: 16.5*ng/40591

cDNA
608801

608801*16.5*"ng"/40591= 248 ng
This was in 1 uL so 248 ng/uL
4-11-13 I currently have 9 uL left
Total amount electroeluted is about 248*"ng"/"uL"*10*"uL"=2480*"ng"

-Approximately how much was there originally (3-26-13)? Lanes 1,2, and 7

were used at this step.
Opened Gel 1 Total 35 cycles
"S:\Research\Cancer_Eradication\Discovering tumor specific antigens\Tumor
cDNA Library\3-27-13 cdna library\gel 1 total 35 cycles 1hr 70V 2p0
exp.jpg"
Cumulative Intensity of 3rd from the top band
(12*"uL"*500*"ng"/(20*"uL")*36*"ng"/(500.0*"ng"))/26894 = 21.6*"ng"/26894

Cumulative intensity lane 1: 859941
859941*21.6*"ng"/26894= 691*"ng" total

Cumulative intensity lane 2: 794374
794374*21.6*"ng"/26894 = 638*"ng" total

Opened Gel 2 total 40 cycles
"S:\Research\Cancer_Eradication\Discovering tumor specific antigens\Tumor
cDNA Library\3-27-13 cdna library\gel 2 total 40 cycles 1hr 70V 2p0
exp.jpg"

Cumulative intensity of 1 kb band: 30047
Conversion factor
(12*"uL"*500*"ng"/(20*"uL")*108*"ng"/(500.0*"ng"))/30047 = 64.8*"ng"/30047

Cumulative intensity lane 7: 517848
517848*64.8*"ng"/30047 = 1117*"ng" total

lanes 1, 2, and 7 were combined so there should have been a total of
691*"ng"+638*"ng"+1117*"ng"= 2446*"ng"

Estimated Percent Recovery: 2480/2446*100.0=101%
So the estimated percent recovery is close to 100%

===========================================================================

Purified Ladder

1 kb band
29683

29683*16.5*"ng"/40591=12.1 ng
The 1 kb band is 108/500=0.216 of all the material so there is a total of
12.1*"ng"/0.216=56.0 ng

so 56.0 ng/uL
There was 10 uL so 56.0*"ng"/"uL"*10*"uL"= 560 ng

I originally (ON 3-22-13) added 30 uL ladder which corresponds to
30*"uL"*500*"ng"/(20*"uL")=750 ng

Estimated Percent Recovery 560/750*100.0 = 74.7%

===========================================================================

Linearized Plasmid (50*"ng"/"uL")

26189

26189*16.5*"ng"/40591= 10.6*"ng"
so 10.6 ng/uL
There was 10 uL so 10.6*"ng"/"uL"*10*"uL=106 ng

I originally (3-22-13) loaded 10*"uL"*50*"ng"/"uL"=500*"ng"

Estimated Percent Recovery = 106/500*100= 21.2%

===========================================================================

Control Insert (40*"ng"/"uL")

30263

30263*16.5*"ng"/40591=12.3*"ng"
12.3 ng/uL
12.3*"ng"/"uL"*10*"uL"=123*"ng"

I originally loaded (3-22-13) 10*"uL"*40*"ng"/"uL" = 400*"ng"

Estimated Percent Recovery = 123/400*100=30.8%