Immunostaining of Formalin-Fixed Paraffin Sections

Five micron formalin-fixed, paraffin embedded sections were cut for immunostaining and analyzed as previously described [63 (link)]. IHC staining was performed at the Pathology Research Core (Mayo Clinic, Rochester, MN) using the Leica Bond RX stainer (Leica). Briefly, tissue slides were dewaxed and retrieved on-line using the following reagents Bond Dewax (Leica) and Epitope Retrieval 2, EDTA based (Leica Biosystems Inc. Buffalo Grove, IL). Tissue slides were retrieved for 30 minutes. The ERβ1 PPG5/10 antibody (Thermo Scientific, Waltham, MA) was diluted 1:75 in Background Reducing Antibody Diluent (Dako, Agilent Technologies, Santa Clara, CA) and incubated for 30 minutes. This antibody has been shown to be highly specific and sensitive for detection of only the full-length form of this receptor in previous IHC studies [59 (link)-61 (link)].

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Reese J.M., Bruinsma E.S., Monroe D.G., Negron V., Suman V.J., Ingle J.N., Goetz M.P, & Hawse J.R. (2017). ERβ inhibits cyclin dependent kinases 1 and 7 in triple negative breast cancer. Oncotarget, 8(57), 96506-96521.

Publication 2017

Bond Dewax Background Reducing Antibody Diluent

Antibody Buffalo Edta Epitope Formalin Paraffin Tissue

Corresponding Organization : Mayo Clinic in Arizona

Other organizations : Mayo Clinic, Mayo Clinic in Florida

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Variable analysis

independent variables

Tissue slides were dewaxed and retrieved on-line using the following reagents Bond Dewax (Leica) and Epitope Retrieval 2, EDTA based (Leica Biosystems Inc. Buffalo Grove, IL)
The ERβ1 PPG5/10 antibody (Thermo Scientific, Waltham, MA) was diluted 1:75 in Background Reducing Antibody Diluent (Dako, Agilent Technologies, Santa Clara, CA) and incubated for 30 minutes

dependent variables

IHC staining results

control variables

Five micron formalin-fixed, paraffin embedded sections were cut for immunostaining
Tissue slides were retrieved for 30 minutes

positive controls

The ERβ1 PPG5/10 antibody has been shown to be highly specific and sensitive for detection of only the full-length form of this receptor in previous IHC studies [59 (link)-61 (link)]

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