» Articles » PMID: 26043998

A Well-based Reverse-phase Protein Array of Formalin-fixed Paraffin-embedded Tissue

Overview
Specialty Molecular Biology
Date 2015 Jun 6
PMID 26043998
Citations 2
Authors
Affiliations
Soon will be listed here.
Abstract

Biomarkers from tissue-based proteomic studies directly contribute to defining disease states as well as promise to improve early detection or provide for further targeted therapeutics. In the clinical setting, tissue samples are preserved as formalin-fixed paraffin-embedded (FFPE) tissue blocks for histological examination. However, proteomic analysis of FFPE tissue is complicated due to the high level of covalently cross-linked proteins arising from formalin fixation. To address these challenges, we developed well-based reverse-phase protein array (RPPA). This approach is a robust protein isolation methodology (29.44 ± 7.8 μg per 1 mm(3) of FFPE tissue) paired with a novel on electrochemiluminescence detection system. Protein samples derived from FFPE tissue by means of laser capture dissection, with as few as 500 shots, demonstrate measurable signal differences for different proteins. The lysates coated to the array plate, dried up and vacuum-sealed, remain stable up to 2 months at room temperature. This methodology is directly applicable to FFPE tissue and presents the direct opportunity of addressing hypothesis within clinical trials and well-annotated clinical tissue repositories.

Citing Articles

Downregulation of antigen presentation-associated pathway proteins is linked to poor outcome in triple-negative breast cancer patient tumors.

Pedersen M, Hood B, Beck H, Conrads T, Ditzel H, Leth-Larsen R Oncoimmunology. 2017; 6(5):e1305531.

PMID: 28638726 PMC: 5468003. DOI: 10.1080/2162402X.2017.1305531.


A melanin-bleaching methodology for molecular and histopathological analysis of formalin-fixed paraffin-embedded tissue.

Chung J, Choi J, Sears J, Ylaya K, Perry C, Choi C Lab Invest. 2016; 96(10):1116-27.

PMID: 27548802 PMC: 7781076. DOI: 10.1038/labinvest.2016.90.

References
1.
Mason J, OLeary T . Effects of formaldehyde fixation on protein secondary structure: a calorimetric and infrared spectroscopic investigation. J Histochem Cytochem. 1991; 39(2):225-9. DOI: 10.1177/39.2.1987266. View

2.
Hultschig C, Kreutzberger J, Seitz H, Konthur Z, Bussow K, Lehrach H . Recent advances of protein microarrays. Curr Opin Chem Biol. 2005; 10(1):4-10. PMC: 7108394. DOI: 10.1016/j.cbpa.2005.12.011. View

3.
Ikeda K, Monden T, Kanoh T, Tsujie M, Izawa H, Haba A . Extraction and analysis of diagnostically useful proteins from formalin-fixed, paraffin-embedded tissue sections. J Histochem Cytochem. 1998; 46(3):397-403. DOI: 10.1177/002215549804600314. View

4.
Chung J, Lee S, Kris Y, Braunschweig T, Traicoff J, Hewitt S . A well-based reverse-phase protein array applicable to extracts from formalin-fixed paraffin-embedded tissue. Proteomics Clin Appl. 2010; 2(10-11):1539-47. PMC: 3777740. DOI: 10.1002/prca.200800005. View

5.
Xie R, Chung J, Ylaya K, Williams R, Guerrero N, Nakatsuka N . Factors influencing the degradation of archival formalin-fixed paraffin-embedded tissue sections. J Histochem Cytochem. 2011; 59(4):356-65. PMC: 3201147. DOI: 10.1369/0022155411398488. View