Fig. 2. Characterization of mAb K56-386. (A) Nesprin-2 pAbK1-immunoprecipitated PAM212 samples were probed with K56-386, which specifically recognizes Nesprin-2 Giant (arrowhead). (B) Equal amounts of HaCaT and PAM212 (mouse) cell lysates were probed with K56-386 and anti-β-tubulin (loading control). (C-G) K56-386 (1:10 dilution) stains the NE in PAM212 keratinocytes (C-E) and in WT mouse-skin frozen sections (F), and is largely absent in KO skin (G). Phalloidin staining delineates the plasma membrane. (G,H) K56-386 (1:10) staining is largely absent in KO epidermis (G); rarely, some residual NE staining is detectable in isolated regions in the KO epidermis, when K56-386 is used undiluted (inset in H). (I-K) The staining pattern of Nesprin-2 Giant is distinct from that of Nesprin-1 Giant in human (I,J) and mouse (K) epidermis. Nesprin-1-Giant localizes to the NE (I, arrowheads) and to cell-cell junctions (I, arrows). Insets, higher-magnifications of boxed areas. epi, epidermis; de, dermis; hf, hair follicle.