Slope/end point analysis of Invader data

• For data retrieval please refer to the manual describing the platforms used to generate the data.

• Paste the data in an empty excel sheet so that cycle numbers start in column A and continue down, the samples starting in column B and onward.

Mark column B, right-click and chose insert. In the inserted column add the time points.

Copy the entire row (A) in the top outlining your wells and paste the whole row under the last data collected. Under this row write the label of each sample.

Usually, only the data range corresponding to up to 10% of the strongest scoring sample is used for each sample. This is chosen as follows. Place the cursor between the brackets in the “10%MAX” box. Delete the x by hitting backspace. Now mark the entire data field and hit enter.

Go back and mark the entire data field again and chose conditional formatting under format. Choose “Cell value is” and “greater than or equal to” and in the third box double click followed by clicking on the 10%MAX value. Then click on format and under color chose red. Click on ok and again ok. All data points higher than 10% of the strongest data point are now marked red.

Now start a slope rate row under your sample definition row. For the first well write “=slope(,)”. Place the cursor before the comma and mark all black data points for that sample. Likewise, place the corresponding time points after the comma. Repeat this for all the samples.

Go to the sheet named slope analysis. This template contains an analysis matrix for duplicate samples and ten known standards.

In the matrix the following are calculated: Average, Standard deviation (SD), Coefficient of variation (CV), Fold over 0, Net signal, Ttest to neighbour and Ttest to 0.

If more than duplicates are run mark the row outlining Average, right click and hit insert. Repeat this for every extra repetition.

If more than 10 samples are analyzed copy the area comprising the values and all calculations and place the curser in the first adjacent empty square. Right click and choose “paste special”. Use the option “formulas” when pasting.

Check all squares containing calculations to make sure that the new repetitions are included and that they all link to the right square. Adjust if necessary.

Now go to raw data and copy the slope rate for one sample. Go back to slope rate analysis tab and mark its position in the matrix. Right click and in “paste special” choose “values”. Click ok. Repeat this for all samples.

Mark the Net signal values and click on the table icon in the toolbar selecting “XY (scatter)” followed by Next.

Choose the tab series and put cursor in the X values box and then mark the copy number series in the matrix.

Click Next and in the following display put in chart title and values for X and Y axis. Click Next followed by finish.

Place cursor over one of the data points and right click followed by “add trendline”. Choose linear in “type” and display R value and equation in “options”.

Use the equation to quantify any unknown samples.

Find the row displaying the time point of interest. Right click the entire row and copy it. Paste the row below, under the row of samples.

Next steps are identical to slope end analysis. Follow instruction from step 3. It is not necessary to use the paste special.

This procedure is identical to slope end analysis (step 4 onwards).