Immunohistochemical Analysis of Tumor Samples

IHC was performed on tissue microarray (TMA) slides with mAbs (1:1000) and developed using the Vectastain Universal kit (Vector Laboratories, Burlingame, CA), as described previously (1 (link)). Results were blindly evaluated by pathologist (HW) and expression levels were categorized as positive or negative (cytoplasmic staining of >10% or <10% of the tumor cells, respectively) and staining intensity as strong, moderate, or no staining. Apoptotic cells were detected in paraffin sections by fluorescence labeled TUNEL staining (Promega, Cat # G3250). Activation of the MapK pathway was evaluated by analysis of the levels of p-ERK (Cell Signaling, Cat # 9160; 1:1000) and the proliferative index of the cancer cells was measured by Ki-67 staining (Thermo Scientific, Cat # RM-9106-S0; 1:200).

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Arumugam T., Deng D., Bover L., Wang H., Logsdon C.D, & Ramachandran V. (2015). New Blocking Antibodies against Novel AGR2-C4.4A Pathway Reduce Growth and Metastasis of Pancreatic Tumors and Increase Survival in Mice. Molecular cancer therapeutics, 14(4), 941-951.

Publication 2015

Vectastain Universal kit TUNEL staining p-ERK

Apoptotic Cancer Cytoplasmic Fluorescence Mabs Microarray Paraffin Pathologist Promega Tissue Tumor Tunel Vector

Corresponding Organization : The University of Texas MD Anderson Cancer Center

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Variable analysis

independent variables

Immunohistochemistry (IHC) performed on tissue microarray (TMA) slides
Monoclonal antibodies (mAbs) used at a dilution of 1:1000

dependent variables

Expression levels of target proteins (categorized as positive or negative based on cytoplasmic staining of >10% or <10% of the tumor cells, respectively)
Staining intensity (categorized as strong, moderate, or no staining)
Apoptotic cells detected by fluorescence labeled TUNEL staining
Activation of the MAPK pathway (evaluated by analysis of the levels of p-ERK)
Proliferative index of the cancer cells (measured by Ki-67 staining)

control variables

Vectastain Universal kit (Vector Laboratories, Burlingame, CA) used for IHC development, as described previously (1 (link))
Evaluation of results by a blinded pathologist (HW)

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

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