Multiple Immunostainings with Different Epitope Retrievals-The FOLGAS Protocol.

We describe a sequential multistaining protocol for immunohistochemistry, immunofluorescence and CyTOF imaging for formalin-fixed, paraffin-embedded specimens (FFPE) in the formalin gas-phase (FOLGAS), enabling sequential multistaining, independent from the primary and secondary antibodies and retrieval. Histomorphologic details are preserved, and crossreactivity and loss of signal intensity are not detectable. Combined with a DAB-based hydrophobic masking of metal-labeled primary antibodies, FOLGAS allows the extended use of CyTOF imaging in FFPE sections.

High Numbers and Densities of PD1 T-Follicular Helper Cells in Triple-Negative Breast Cancer Draining Lymph Nodes Are Associated with Lower Survival.

Breast cancer tumor draining lymph nodes (TDLNs) display distinct morphologic changes depending on the breast cancer subtype. For triple-negative breast cancers (TNBC), draining LNs display a higher amount of secondary lymphoid follicles, which can be regarded as a surrogate marker for an activated humoral immune response. In the present study, we focus on PD1 T-follicular helper cells (Tfh) in TDLNs of TNBC, since PD1 Tfh are drivers of the germinal center (GC) reaction.