Supplemental Material

This article contains the following supporting material:

• Figure 1 - Steady-state localization of mature Pmel is not significantly altered in melan-si-1 compared with melan-a.
  IFM analysis of melan-a (A-C, G-J) and melan-si-1 (D-F, K-M) cells. Mouse melanocytes were colabeled with HMB-45 (anti-mature Pmel; A, D, G, K) and either TA99 (anti-Tyrp1; B, E) or 1D4B (anti-LAMP1; H, L), followed by Alexa-594 and –488-conjugated isotype- or species-specific secondary antibodies. Cells were analyzed by IFM, and z-series images were deconvolved. C, F, J, M; corresponding bright field images. Insets within bright field panels, 5X magnifications of indicated regions with red and green channels merged. To better visualize any coincidence of labeling of HMB-45 with markers of both mature melanosomes (Tyrp1) and late endosomes/ lysosomes (LAMP1), fluorescence intensity levels for melan-si-1 images were increased disproportionately to those of melan-a. Bar, 20 μm.

  Supplementary Materials and Methods
  Rat monoclonal antibody 1D4B to mouse LAMP1 was obtained from Developmental Studies Hybridoma Bank (Iowa City, IA).