Cancer Diagnosis Program | Biorepositories and Biospecimen Research Branch

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One-step preservation of phosphoproteins and tissue morphology at room temperature for diagnostic and research specimens.

Author(s): Mueller C, Edmiston KH, Carpenter C, Gaffney E, Ryan C, Ward R, White S, Memeo L, Colarossi C, Petricoin EF 3rd, Liotta LA, Espina V

Publication: PLoS One, 2011, Vol. 6, Page e23780

PubMed ID: 21858221 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 fixation duration and specimen size on levels of phosphorylated and non-phosphorylated proteins in colon mucosa from a single patient and to compare levels between the colon mucosa of 2 patients. Units of size were not specified but all specimens were <1 cm^3.

   Summary of Findings:

   BHP fixation for 7 days instead of 48 h had no effect on the levels of any of the proteins examined by reverse phase microarray (11 phosphorylated, 2 cleaved and 10 non-phosphorylated proteins). Further, BHP-fixation for 3 months had no effect on levels of the two proteins investigated by immunohistochemistry (phosphorylated extracellular signal-regulated kinases (ERK) or cytokeratin 20). Tissue size had no effect on the levels of all 11 phosphorylated proteins measured. Levels of all 19 phosphorylated and non-phosphorylated proteins were comparable between the BHP-fixed colon mucosa from 2 patients with similar processing times.

   Biospecimens

   • Tissue - Colorectal

   Preservative Types

   • Other Preservative

   Diagnoses:

   • Not specified

   Platform:

   Analyte	Technology Platform
   Protein	Reverse phase protein microarray
   Protein	Immunohistochemistry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Preservation	Time in fixative	48 h 7 days 3 months
   Biospecimen Aliquots and Components	Aliquot size/volume	0.5-2.5 x 10^7 (unspecified units)
   Immunohistochemistry Specific	Targeted peptide/protein	Cytokeratin 20 ERK

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2. Study Purpose

   The purpose of this study was to compare tyrosine phosphatase activity, histological preservation and immunohistochemical staining between FFPE and BHP-fixed specimens. The study included an unspecified number of breast, uterine, colon, lymph node and prostate specimens from patients undergoing surgery. The effect of fixation duration was investigated using an tyrosine phosphatase activity ELISA with fixed but not embedded breast tissue.

   Summary of Findings:

   In breast specimens, tyrosine phosphatase activity declined with increasing fixation duration in BHP or formalin, but activity levels were higher in formalin fixed specimens than BHP-fixed specimens at all time-points. BHP fixation led to better nuclear structure preservation and less nuclear shrinkage than formalin fixation. Color fidelity, cell size, and overall cell structure were comparable between BHP-fixed and FFPE specimens. IHC staining of 13 of 14 non-phosphorylated proteins and all 6 phosphorylated proteins was comparable to or better in BHP-fixed specimens than FFPE specimens, but staining for caudal-type homeobox (CDX2) was slightly weaker in FFPE specimens compared to BHP-fixed specimens.

   Biospecimens

   • Tissue - Breast
   • Tissue - Uterus
   • Tissue - Prostate
   • Tissue - Lymph Node
   • Tissue - Colorectal

   Preservative Types

   • Formalin
   • Other Preservative

   Diagnoses:

   • Not specified

   Platform:

   Analyte	Technology Platform
   Morphology	H-and-E microscopy
   Protein	Immunohistochemistry
   Protein	ELISA

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Preservation	Type of fixation/preservation	Biomarker and histology preservative (BHP) Formalin (buffered)
   Immunohistochemistry Specific	Targeted peptide/protein	CDX2
   ELISA Specific	Targeted peptide/protein	Tyrosine phophatase activity
   Biospecimen Preservation	Time in fixative	1 min 5 min 15 min

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