Protocol Modifications Influence the result of EGF receptor immunodetection by EGFR pharmDx in Paraffin-Embedded Cancer Tissues

Author(s): Derecskei Katalin, Moldvay Judit, Bogos Krisztina, Timar Jozef

Publication: Pathol Oncol Res, 2006, Vol. 12, Page 243

PubMed ID: 17189989 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The aim of this study was to complete a comparative analysis of appropriate EGFR immunostaining of lung carcinoma specimens after incubation with the primary antibody for 30 min at room temperature or overnight at 4 degrees C.

   Summary of Findings:

   Results using the EGFR pharmDx kit and protocols yielded seven EGFR immuno-negative cases of the fifty examined. Extended incubation with the primary antibody overnight at 4 degrees C increased both the number of positively stained cells as well as the number of immuno-positive cases (42 vs 46).

   Biospecimens

   • Tissue - Lung

   Preservative Types

   • Formalin

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   Protein	Immunohistochemistry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Incubation duration/condition	Overnight at 4 degrees C 30 min at room temperature

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2. Study Purpose

   Comparative analysis of appropriate EGFR immunostaining using the FDA approved detection method (dextran polymer conjugate) with an alternative detection method using the LSAB kit.

   Summary of Findings:

   EGFR immunostaining was unaffected by the alternative method of detection (LSAB kit) in comparison to the FDA approved detection method (dextran polymer conjugate).

   Biospecimens

   • Tissue - Lung

   Preservative Types

   • Formalin

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   Protein	Immunohistochemistry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Immunohistochemistry Specific	Detection method	Dextran polymer conjugate LSAB kit

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3. Study Purpose

   The purpose of this study was to conduct a comparative analysis of appropriate EGFR immunostaining of lung carcinoma specimens using the FDA approved antigen retrieval technique for EGFR detection (proteinase K treatment) and the alternative method of microwave heating in citrate buffer.

   Summary of Findings:

   Microwave heating in citrate buffer resulted in an increase in the percentage of EGFR positively stained cells when used in combination with prolonged primary antibody incubation when compared to room temperature proteinase K digestion and a 30 min primary antibody incubation (> 50% versus <10%, respectively).

   Biospecimens

   • Tissue - Lung

   Preservative Types

   • Formalin

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   Protein	Immunohistochemistry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Temperature of heat-induced retrieval	Room temperature Microwave heating
   Analyte Extraction and Purification	Incubation duration/condition	30 min at room temperature Overnight at 4 degrees C
   Analyte Extraction and Purification	Antigen retrieval	0.05M citrate buffer 0.1% proteinase K digestion

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