The extraorbital and intraorbital glands were extracted and fixed in optimal cutting temperature compound (Sakura Finetek USA, Inc., Torrance, CA). The glands were preserved at -80°C until sectioning. The sections were cut at 4 μm and subjected to hematoxylin-eosin staining and immunostaining for GFP, CD3, CD11b, and CD19. The primary antibodies used were Alexa Fluor® 488 anti-GFP antibody, anti-mouse Alexa Fluor® 700 CD3 antibody, Alexa Fluor® 594 anti-mouse CD11b antibody, and Alexa Fluor® 647 anti-mouse CD19 antibody (all from BioLegend®, San Diego, CA). A mounting medium containing DAPI (VECTASHIELD® Mounting Medium, Vector Laboratories, Inc., Burlingame, CA) was used for counterstaining.
For TUNEL staining, ApopTag® Red in situ Apoptosis Detection Kit (EMD Millipore, Billerica, MA) was used.
To detect MSCs in the glands, the sections were stained with mouse anti-human nuclei (Cy3 conjugate, 1:50) (MAB1281C3, Millipore, Billerica, MA) and mouse anti-human mitochondria (1:50) (MAB1273, Millipore), followed by Alexa Fluor 488 goat anti-mouse IgG (1:500) (A11001, Molecular Probes®/Life Technologies, Eugene, OR).
For TUNEL staining, ApopTag® Red in situ Apoptosis Detection Kit (EMD Millipore, Billerica, MA) was used.
To detect MSCs in the glands, the sections were stained with mouse anti-human nuclei (Cy3 conjugate, 1:50) (MAB1281C3, Millipore, Billerica, MA) and mouse anti-human mitochondria (1:50) (MAB1273, Millipore), followed by Alexa Fluor 488 goat anti-mouse IgG (1:500) (A11001, Molecular Probes®/Life Technologies, Eugene, OR).