Fig. 3. Role of immunoglobulins in efficiency of C3 deposition on clinically approved liposomes:

a) schematic representation of LipoDox^® (a generic version of Doxil^®, a regulatory approved liposomal doxorubicin); b) effect of immunoglobulin depletion and reconstitution in plasma samples of healthy donors. These experiments were repeated independently 3 times in 3 different plasma samples with similar results; c) effect of immunoglobulin depletion and reconstitution in healthy sera (n=4 subjects), measured with dot blot. Each dot represents the mean of 3 technical replicates per sample; d) correlation between levels of immunoglobulin and C3 bound to LipoDox^® in healthy sera (n=12 subjects). Red points indicate male samples and blue points indicate female samples. Each point is the mean of 3 experimental and 3 technical replicates per sample; e) nanoparticles were incubated in serum with EGTA/Mg²⁺ or EDTA (n=5 subjects). In panels (c) and (e) same colour refers to the same individual. Each dot represents the mean of 3 technical replicates per sample; f) immunoglobulin depletion and reconstitution in lepirudin-anticoagulated plasma from breast cancer patients (n=8 subjects). Each dot represents the mean of 3 technical replicates per sample. EDTA-anticoagulated plasma was from the same patient; g) schematic representation of Onivyde^®, a clinically approved liposomal irinotecan; h) effect of immunoglobulin depletion and reconstitution in plasma of healthy donors. These experiments were repeated independently 3 times in 3 different plasma samples with similar results; i) effect of immunoglobulin depletion and reconstitution in healthy sera (n=3 subjects). Each dot represents the mean of 3 technical replicates per sample; j) association between levels of immunoglobulin and C3 bound to Onivyde^® in healthy sera (n=12 subjects). Red points indicate male samples and blue points indicate female samples. Each point is the mean of 3 experimental and 3 technical replicates per sample; k) effect of EGTA/Mg²⁺ or EDTA on C3 deposition in healthy sera (n=5 subjects). In panels (i) and (k) the same colour refers to the same individual. Each dot represents the mean of 3 technical replicates per sample; l) IgG depletion and reconstitution in lepirudin-anticoagulated plasma from breast cancer patients (n=8 subjects). Each dot represents the mean of 3 technical replicates per sample. EDTA-anticoagulated plasma was from the same patient. In all graphs, lines connecting points indicate measures from the same subject. Colours designate different individuals. Statistical analysis is explained in Methods