Effects of Decalcifying Agents of Variable Duration on PD-L1 Immunohistochemistry

Am J Clin Pathol. 2020 Jan 2;153(2):258-265. doi: 10.1093/ajcp/aqz161.

Abstract

Objectives: To evaluate the effects of decalcifying agents on programmed cell death ligand 1 (PD-L1) immunohistochemistry (IHC).

Methods: Fragments of 10 placentas (high PD-L1 expressor) and 10 lungs (lower PD-L1 expressor) were formalin-fixed and subjected to four decalcifying solutions (EDTA, formic acid/MasterCal IM Plus [FA/MC], 12% HCl, and Decal STAT/23% HCl) for 1, 2, 6, or 24 hours. H&E staining and PD-L1 using IHC 22C3 pharmDx were performed, and PD-L1 staining was assessed.

Results: Minimal to no change in staining intensity or proportion of stained cells was seen with EDTA or FA/MC at all decalcifying durations. Both HCl-based decalcifiers demonstrated a progressive decrease in percentage of positive cells and staining intensity with longer decalcifying duration, particularly with Decal STAT.

Conclusions: EDTA and FA/MC have little effect on PD-L1 expression. 12% HCl causes a progressive decline in staining. Decal STAT dramatically reduced staining with all treatment durations, especially at 24 hours.

Keywords: Decalcification; IHC; PD-L1.

MeSH terms

  • B7-H1 Antigen / analysis*
  • Calcium Chelating Agents / pharmacology*
  • Edetic Acid / pharmacology*
  • Female
  • Formates / pharmacology
  • Humans
  • Hydrochloric Acid / pharmacology*
  • Immunohistochemistry
  • Lung / chemistry
  • Placenta / chemistry
  • Pregnancy
  • Prospective Studies

Substances

  • B7-H1 Antigen
  • CD274 protein, human
  • Calcium Chelating Agents
  • Formates
  • formic acid
  • Edetic Acid
  • Hydrochloric Acid