Fig. 7

Fluorescent antibody labeling of membrane proteins in fixed cells and tissues in mouse and human. a Representative gating strategy of one experiment analyzed by flow cytometry with cryopreserved and cryopreserved+fixed PBMCs from healthy donors (n  =  3). PBMCs were stained with anti-human CD45, CD3, CD19, CD4, and CD8 monoclonal antibodies (mAbs). T cells were selected by the positive expression of CD3, whereas B cells were selected from the CD3-negative fraction (Non T cells) and by the positive expression of CD19. CD4-positive and CD8-positive T cells were selected from CD3-positive T cells. Box plots show the percentage of positive cells in each subpopulation for cryo (blue) and cryo+fixed (orange) PBMCs analyzed by flow cytometry. b Representative histograms of the mean fluorescent intensity (MFI) with cryopreserved and cryopreserved+fixed PBMCs from healthy donors (n = 3) stained with anti-human CD45, CD3, CD19, CD4, and CD8 mAbs. Bar plots show the MFI expression for three fresh and fixed PBMC samples analyzed by flow cytometry. c, d Representative histograms of MFI of fresh and fixed PBMCs from healthy donors (n = 4) stained with anti-human CD45, CD3, CD4, and CD8 mAbs, anti-β2M, anti-CD298, and LMOs analyzed by flow cytometry. f Representative histograms of the MFI from cryopreserved (blue) and cryopreserved+fixed (orange) human colon samples processed and stained with anti-human CD45, CD3, CD11b, and EpCAM mAbs and analyzed by flow cytometry. g Multiplex fluorescence tissue imaging of a human prostate cancer section, DSP-fixed (top) or formalin-fixed (bottom) paraffin-embedded, captured using Phenocycler. Images show hematoxylin and eosin staining, five-color overlay, and individual SMA, Pan-CK, E-cadherin, and p63 antibody staining