Quantitative assessment shows loss of antigenic epitopes as a function of pre-analytic variables.

Author(s): Bai Y, Tolles J, Cheng H, Siddiqui S, Gopinath A, Pectasides E, Camp RL, Rimm DL, Molinaro AM

Publication: Lab Invest, 2011, Vol. 91, Page 1253-61

PubMed ID: 21519325 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of cold ischemia time, surgical procedure type, and the analysis of tissue microarrays versus whole tissue sections on quantitative immunofluorescence of proteins and phospho-proteins using AQUA. The median time between acquisition of core needle biopsy and surgical resection was 27.5 days.

   Summary of Findings:

   When a cohort of 20 paired CNB and SR were examined in a tissue microarray format (AQUA scores from 2 spots for each type of specimen were averaged), no statistically significant differences were seen in the antigenicity of pERK, pAKT, pTyr, ER, p53, or Ki67. However, insignificant decreases in pERK, pAKT, pTyr, and Ki67 were seen for SR compared with CNB. When whole tissue sections were evaluated from a second cohort of 14 matched CNB and SR, pAKT, pERK, ER, cytokeratin, and pS6K1 immunofluorescence was significantly lower in SR (average of 19 20x fields of view) compared to CNB (average of 11 20x fields of view; p<0.05). For ER, the authors report that none of the cases would have changed from an ER-positive status in the CNB to an ER-negative status with the SR. At the same time, total AKT, ERK, S6K1, Ki67, and GAPDH were not affected by surgical procedure type, however the authors subsequently determined their sample size was not large enough to detect total protein epitope differences for AKT, ERK, S6K1, Ki67, and GAPDH.

   Biospecimens

   • Tissue - Breast

   Preservative Types

   • Formalin

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   Protein	Immunohistochemistry
   Protein	Tissue microarray
   Small molecule	Immunohistochemistry
   Morphology	Fluorescent microscopy

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Acquisition	Cold ischemia time	0 min 1-5 h
   Immunohistochemistry Specific	Targeted peptide/protein	pAKT AKT pERK ERK pS6K1 S6K1 GAPDH Ki67 ER pTyr p53 Cytokeratin
   Biospecimen Aliquots and Components	Type of slide	Tissue microarray Whole tissue section
   Biospecimen Aliquots and Components	Aliquot size/volume	Two 1.5 mM tissue microarray cores An average of 11 20x fields of view An average of 19 20x fields of view
   Biospecimen Acquisition	Method of tissue acquisition	Core needle biopsy Surgical resection

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