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. 2012 Sep 4;7(9):e44505. doi: 10.1371/journal.pone.0044505

Figure 4. Betaine increases solubility of intracellular FVIII.

Figure 4

CHO cells expressing eGFP-tagged FVIII-BDD protein were incubated with and without betaine. (A,B) Flow cytometry analysis was used to determine the eGFP-signal in untreated cells versus cells treated with betaine or control substance BHA. The mean eGFP intensity (X mean GFP) in the range M1 was used as distinctive parameter. (A) representative histogram after 48 h of treatment and (B) values after 72 h presented as means ± SEM of 3 independent experiments. ANOVA **P<.001. (C,D) After 72 h incubation cells were successively lysed in PBS/0.5% Triton X-100 and PBS/1% SDS. (C) FVIII antigen was determined in both fractions by indirect ELISA. (D) Triton X-100-soluble and insoluble fractions were separated on SDS-polyacrylamid gradient gels, and hFVIII light chains (lc), eGFP in hFVIII-single chain (sc) and GAPDH were detected by Western blot. Δ indicates lower band of hFVIII lc doublet.