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Table 1 Characteristics for immunostaining and scoring

From: Presence of HER4 associates with increased sensitivity to Herceptin™ in patients with metastatic breast cancer

Protein

HER2 EC

phospho-HER2 (pY1248)

p27Kip1

Oestrogen receptor α

Progesterone receptor

Antibody

Rabbit mAb

Mouse mAb

Mouse mAb

Mouse mAb

Mouse mAb

Origin

Thermo Scientific

Thermo Scientific

Lab Vision

Novocastra

Novocastra

Clone

SP3

PN2A, Ab-18

DCS-72.F6, Ab-1

6F11

1A6

Dilution of primary antibody

1:50

1:100

1:100

1:60

1:40

Staining pattern

Membrane

Nucleus and cytoplasm

Nucleus

Epitope retrieval

Heat induced, 10 mM citric acid buffer, pH 7.3

Heat induced, 10 mM sodium-citric acid pH 6.0

Heat induced, 10 mM citric acid buffer, pH 7.3

Blocking

Endogenous peroxidase blocking

Primary antibody

30 min, room temperature

Overnight, 4°C

30 min, room temperature

60 min, room temperature

 

Detection system

EnVision™ Dual Link System; DAB + chromogenic substrate

iVIEW™ DAB Detection Kit

 

Scoring in accordance with

HercepTest guidelines

Immunoreactive score

Manufacturer's guidelines

  1. Anti-HER1 to 4 and anti-Ki-67 antibodies were applied as described previously [27].
  2. EC = antibody binds to extracellular domain; HER = human epidermal growth factor receptor; mAb = monoclonal antibody.