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I’m aware of how to get around the problem, I could extend the previous post. The actual problem is that any changes that I make to the code are now lost as soon as I close and reopen the project.


I figured it out thanks to some help from a colleague. The problem was that the project was set to use the B&W color space. The setting is under ‘Image -> Color Settings -> Convert to Color’ (in the image below it is B&W Color, but I wanted to use RGB). The setting was not changed with the code changes, only the project was. The solution was to change the project setting back to RGB from RGB/CMYK.

.a. 10 min) were incubated with anti-activated NF-κB (p65) antibody (1:1,000 dilution) at 4 °C overnight. The membranes were subsequently washed with PBS three times and incubated with horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (1:10,000 dilution) at 37 °C for 1 h. After further washes, the membrane was subjected to chemiluminescence-based detection (Millipore, USA), and the band intensity was quantified using the ImageJ software (NIH).

Statistical analysis {#Sec23}

The data were presented as mean ± standard deviation from three independent experiments, unless stated otherwise. Statistical analysis was performed using either Student’s t test or one-way analysis of variance followed by Tukey’s post hoc test.

Additional files {#Sec24}

Additional file 1:**Figure S1.** Berberine induces HUVECs apoptosis in a time-dependent manner. HUVECs were treated with berberine (10 μM) for the indicated periods of time. Cell apoptosis was analyzed by flow cytometry. The data are shown as mean ± SD (*n* = 3). \* *P* \

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