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In total, the treatment usually takes around 1015 minutes, but Daniel's took much longer because 1

2.5 l of this cell suspension was mixed with 7.5 l of lysis buffer [200 mM Tris-HCl (pH 7.5), 50 mM EDTA, 0.5% (w/v) SDS] directly on a slide and incubated for 4 min followed by tilting the slides to 30 causing the drops to run down by gravity

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Over the past week emails have been exchanged several time a day with the participants all having an input into the content of the emerging guideline

Possible causes: dead space accumulation (discussed above), inaccurate water volume at reconstitution (more water means more dilute, means more volume per dose, means fewer doses from the same amount of peptide), or air in the vial displacing solution