Prefoldin enhances the efficiency of TRiC-mediated µ13σ33 heterohexamer assembly. (A) Workflow for µ13σ33 heterohexamer assembly assay. (B) Native-immunoblot of µ13σ33 assembly reactions conducted in the presence of BSA or PFD. +,++, 1:1 or 4:1 PFD/BSA:TRiC molar ratio. (C) Quantification of μ1/σ3 assembly abundance from B. (D) Workflow for assembly assay with denatured 35S-labeled σ3 and buffer, PFD, or BSA preincubation. (E) Native-PAGE of µ13σ33 assembly reactions conducted with buffer, PFD, or BSA. (F) Quantification of μ1/σ3 assembly abundance from E. (G) Workflow for PFD time-of-addition µ13σ33 heterohexamer assembly assay. (H) Native-immunoblot of µ13σ33 assembly reactions using independent PFD conditions. (I) Quantification of μ1/σ3 assembly abundance from H. For all quantifications, data are plotted as mean ± SD for three independent replicates (*P < 0.05; **P < 0.01; ***P < 0.001; unpaired two-tailed t test). Asterisk corresponds to aggregated σ3 and the arrowhead corresponds to TRiC/σ3.