Interaction of tagged NBEAL2 and SEC22B in HEK293 cells, and native proteins in MKs. (A) HA-tagged protein was immunoprecipitated from lysates of HEK293 cells expressing both HA-tagged NBEAL2 (HA-NBEAL2) and FLAG-tagged SEC22B (FLAG-SEC22B). Immunoblots of lysate (input) and immunoprecipitates probed with anti-FLAG and anti-HA antibodies show HA-NBEAL2 pulled down FLAG-SEC22B. (B) IP of FLAG-tagged protein showed that FLAG-SEC22B pulled down HA-NBEAL2. (C) Lysate from primary human MKs was immunoprecipitated with anti-SEC22B antibody; immunoblot analysis shows that endogenously expressed SEC22B pulled down native NBEAL2. (D) IP of NBEAL2 from primary human MK lysate showed that NBEAL2 pulled down SEC22B. (E) Fixed primary human MKs immunostained for NBEAL2 (green) and SEC22B (red) were imaged via laser fluorescence confocal microscopy. XY profiles of midcell z sections from a representative mature cell show substantial overlap of NBEAL2 and SEC22B (bars = 10 μm). Images were obtained with an oil immersion objective (60×/1.35 NA) using a Quorum spinning disc confocal inverted fluorescence microscope. DAPI, 4′,6-diamidino-2-phenylindole; IgG, immunoglobulin G.