Generation and characterization of the anti-mutated CALR antibody. In panel a, the sequence of the 17-mer peptide used for generation of the anti-mutated CALR antibody is shown in relation with the amino-acid sequence of wild-type CALR and the predicted sequence of mutated CALR originated from the del52 and ins5 abnormalities. In panel b, gel electrophoresis of the mutated calreticulin prepared in BL21DE3RIPL (lanes 2 and 3) and JM109DE3 (lanes 4 and 5) E. Coli strains; only in the latter strain successful production of calreticulin was obtained. Calreticulin has a molecular weight of 47 kDa, but migrates in these conditions at an apparent higher molecular weight likely owing to a specific conformational pattern. In panel c, western blot analysis shows anti-mutated CALR antibody selectivity for mutated protein vs the wild-type form. CALR recombinant protein expressed in E. Coli was used as a positive control. GAPDH was used as a loading control.