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As previously described, regions of interest were drawn on the vascular wall to determine calcium signal from smooth muscle cells and background was substracted using MetaFluor (Universal Imaging Software) [17].
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The background was substracted by using the negative isogenic MC4100 Hfq− strain.
Background was substracted from raw data and infected cells were normalized to 100%.
Subsequently, local background was substracted and raw data were normalized and t-test performed for three replicates.
Capacitances were compensated and leak currents were substracted (P/n) using the facilities of the Pulse software (HEKA Elektronik, Lambrecht, Germany).
The intensity of the background surrounding each spot was substracted from that of each of the spots.
Enzymatic activity was measured in the linear part of the curve (5 20 min) as the rate of the curve, to which background activity of uninfiltrated WT tissue was substracted.
The spectrum for buffer alone was substracted from the measurements and the data analysed and visualised using Microsoft Excel.
Background activity of sample control without D-galacturonic acid was substracted from sample measurements with D-galacturonic acid.
The initial scattered light intensity (I0) was mainly attributed to factors such as the media background or the type of the MTP and thus was substracted from the residual scattered light data (I-I0) [ 30] or fluorescence data (where appropiate).
Emission spectra were collected in the 300 400 nm region (with 2 nm-increments), with blanks substracted, using a black flat-bottom, low-binding 96-well microplate (Greiner Bio-one).
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