Characterization of anti-TIMP-1 monoclonal antibodies for immunohistochemical localization in formalin-fixed, paraffin-embedded tissue.
The aim of this study was to evaluate seven anti-TIMP-1 (tissue inhibitor of metalloproteinase-1) monoclonal antibodies by immunohistochemical (IHC) staining of formalin-fixed, paraffin-embedded (FFPE) tissue. Detection of the TIMP-1 protein was studied by IHC in FFPE human archival normal and neoplastic samples. Indirect IHC technique was used, and the seven antibodies (clones VT1, VT2, VT4, VT5, VT6, VT7, and VT8) were tested in various concentrations using different pretreatment protocols. All seven VT antibodies specifically immunostained the cytoplasm of islets of Langerhans cells in normal pancreas, epithelial cells of hyperplastic prostate, tumor cells of medullary thyroid carcinoma, and fibroblast-like cells of malignant melanoma. Specificity of the anti-TIMP-1 antibodies was confirmed by several controls, e.g., Western blotting on proteins extracted from FFPE tissue showed that the VT7 antibody reacted specifically with a protein band of approximately 28 kDa, corresponding to the molecular mass of TIMP-1. However, sensitivity varied with the different antibodies. Use of heat-induced epitope retrieval (HIER) and the VT7 clone applied at low concentrations demonstrated more intense immunoreactivity with the TIMP-1-positive cell types compared to the other six clones. Furthermore, when tested on a range of normal and neoplastic endocrine tissues, the VT7 clone demonstrated immunoreactivity with all neuroendocrine cell types. In conclusion, all seven antibodies detected TIMP-1 protein in various normal and neoplastic FFPE tissues, but one clone, VT7, was superior for IHC staining of TIMP-1 in FFPE tissue sections when using HIER.
Sorensen
Irene Vejgaard
IV
Institute of Veterinary Pathobiology, The Royal Veterinary- and Agricultural University, Ridebanevej 9, 1870 Frederiksberg, Denmark.
Fenger
Claus
C
Winther
Henrik
H
Foged
Niels T
NT
Lademann
Ulrik
U
Brünner
Nils
N
Usher
Pernille A
PA
D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D005404:Fixatives; D005557:Formaldehyde; D006801:Humans; D007150:Immunohistochemistry; D009369:Neoplasms / Q000378:metabolism; D009490:Neurosecretory Systems / Q000378:metabolism; D016612:Paraffin Embedding; D012680:Sensitivity and Specificity; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000276:immunology*
D000911:Antibodies, Monoclonal; D005404:Fixatives; D019715:Tissue Inhibitor of Metalloproteinase-1; D005557:Formaldehyde