Table 3.

Antibodies used for characterization of the sections prior to tetramer staining

AntigenPrimary antibodyDetection antibody
InsulinPolyclonal guinea pig anti-insulin (Dako; 1/140 dilution, 1-h incubation)Polyclonal goat anti–guinea pig IgG, highly cross-adsorbed, Alexa Fluor 488 (Invitrogen; 1/1000 dilution, 30-min incubation)
HLA-ABCMouse monoclonal (clone W6/32) IgG2a against a monomorphic epitope on the 45 kD polypeptide products of the HLA-A, B and C loci (Dako; 1/100 dilution, 1 h incubation)Polyclonal goat anti–mouse IgG2a, isotype-specific, Alexa Fluor 594 (Invitrogen; 1/1000 dilution, 30-min incubation)
CD8Mouse monoclonal (clone HIT8a) IgG1 against the CD8 alpha subunit (BD; 1/100 dilution, 1-h incubation)F(ab’)2 fragment of goat anti-mouse IgG, Alexa Fluor 594 (Invitrogen; 1/1000 dilution, 30-min incubation)