Format of Roche LightCycler output files *.ABT / *.CCC for colour compensation
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6.7 years ago
me • 0

I am automating a workflow for analysis of real-time PCR data sourced from a Roche LightCycler. There are multiple channels for sensing different fluorophores that have a slightly overlapping fluorescence spectrum as detailed here (referred to as "crosstalk"). The detected signals are thus a linear weighting of the true fluorescence, as detailed in Table 1 / Formula 1 here. This is corrected with colour compensation as detailed in the first link.

There is very little (i.e. no) information regarding the format of the older LightCycler file formats ABT (details a particular run) and CCC (details the colour compensation), and I was hoping someone could please provide guidance. 

In the ABT file there is a section ^CCC (all section headers appear to start with ^) which includes the location of the colour compensation file and its contents (a 12x4 matrix of numbers that I am trying to make sense of). In my case they appear as (with file paths changed):

1	1	1
-1.33786041651041E-8	3.39600416861281E-6	-0.000392286209994558	0.196738637483626
-6.43980805238762E-9	1.50246180493155E-6	-0.000107523827209977	0.112676603581708
-2.23359269130001E-8	4.9982365413899E-6	-0.000398780255441468	0.0600920934768699
1.55891872931679E-5	-0.00505090913467267	0.162405918150517	34.8790113064609
2.27218464185121E-8	-0.000146878157270101	0.0123033869019081	1.42038161205974
-1.42030623826416E-7	1.40575424253722E-6	0.0012288371257375	0.290852973511342
2.1866259759953E-9	9.8947181762574E-7	-0.00031255171342293	0.199654709634488
-1.40190153175352E-5	0.00289266217140399	-0.188211203807182	5.67986350485893
-4.39402518213953E-6	0.000909318177116326	-0.0584074241424017	1.73152377696672
7.01531463682071E-7	-0.000117467670542772	0.0039676094304943	0.282456463232278
1.37437444939199E-7	-2.4255075537722E-5	0.000917545927808637	0.132638408109113
1.31314492531558E-5	-0.00255990120176339	0.143275086483057	-1.36050870155249

What do these numbers describe?

EDIT: Since originally posting, I accessed a colour compensation file and found that it is identical to the numbers in the ABT.

qPCR crosstalk • 1.8k views

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