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Table 2 Antibodies used for immunohistochemistry and EIA; GdC15, GdQ13 and GdN20 were purchased from SantaCruz Biotechnologies, Santa Cruz, CA (SCBT)

From: Glycodelin A is a prognostic marker to predict poor outcome in advanced stage ovarian cancer patients

Immunohistochemistry
Antibody Epitope Source/ Clonality Epitope retrieval Blocking Dilution Incubation Reaction system used for detection
GdC15 (SCBT) C-terminal goat/polyclonal Proteinase K (30 min, RT) UV-Block (45 min) 1:1000 (UV Block) 30 min (RT) Vectastain elite kit (goat IgG)
GdQ13 (SCBT) C-terminal goat/polyclonal citrate buffer (pH 6, 5 min, pressure cooker) UV-Block (45 min) 1: 300 (UV Block) o.n. (4°C) Vectastain elite kit (goat IgG)
GdA (Jeschke et al. 2006) Mixed glycan/protein epitope mouse/ monoclonal citrate buffer (pH 6, 5 min, pressure cooker) 1.5% horse serum (20 min) 1:3000 (DAKO dil.) o.n. (4°C) Vectastain elite kit (mouse IgG)
Competitive enzyme immuno assay (EIA)
Antibody Epitope Source/ Clonality Competing peptide Blocking Dilution Incubation Reaction system used for detection
GdQ13 (SCBT) C-terminal goat/ polyclonal GdQ13-P (SCBT) 1% BSA, 0.5% NaN3 in PBS 1.5 ng/ml 1 hour (RT) POD-strepavidin
GdN20 (SCBT) N-terminal goat/ polyclonal GdN20-P (SCBT) 1% BSA, 0.5% NaN3 in PBS 6 ng/ml 1 hour (RT) POD-strepavidin
  1. rt = room temperature, BSA = bovine serum albumin, PBS = phosphate buffered salt solution, o.n. = over night, POD = horse radish peroxidase.
  2. Competing peptides came from SCBT as well and were biotinylated by the manufacturer; unmodiefied GdQ13-P, GdN20-P were used for setting up the standard curve. Preparation of GdA is published in Jeschke et al. 2006. Epitope retrieval: Proteinase K was from Quiagen (Hilden, Germany), citrate buffer contained 0.1 M citric acid and 0.1 M sodium citrate in distilled water (pH 6.0). Blocking: UV-Block was purchased from Thermo Scientific (Bonn, Germany) and horse serum was a component of the Vectastain elite kit (Vector Laboratories, Burlingame, USA).