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The upper phase was discarded and to the lower phenolic phase an equal volume of extraction buffer was added, and the mixture was incubated on ice for 30 min with shaking at 150 rpm

(3) D Wensink, M A E M Wagenmakers, and J G Langendonk (2021) Afamelanotide for prevention of phototoxicity in erythropoietic protoporphyria Expert Review of Clinical Pharmacology, 2021 Feb, Volume 14 (Issue 2), Pages 151-160

However, as observed under a transmission electron microscope, IBs were still evident (Additional file 2: Fig