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All cells were fixed, permeabilized and washed as per protocol.
We included in our experiment cells with neither TdT or F-dUTP (autofluorescence control), cells with F-dUTP but no TdT (negative control), cells with TdT and F-dUTP (experimental control) and a positive control, cells incubated for 10 min at RT with 0.5 mg/ml DNaseI before addition of TdT and F-dUTP.
Flow produced histograms of log green fluorescence.
The experimental controls gave a peak well to the right of autofluoresence controls, and the positive controls gave a peak further right again (a little too clean perhaps). However the negative control, which I would expect to overlay the experimental control appeared somewhere between experimental and positive control.
This unusual negative control situation prevents us from making confident interpretations of our other findings.
Any explanations for the negative control position?
Is this a reasonable positive control?
Your assistance is again greatly appreciated.
Justin Lamb
Cell Pathology Laboratory
University Medical School
Aberdeen
Scotland
j.lamb@abdn.ac.uk
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