Cardiovascular Pathology
Volume 19, Issue 3 , Pages 136-146, May 2010

Creation, validation, and quantitative analysis of protein expression in vascular tissue microarrays

  • Marc K. Halushka

      Affiliations

    • Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD, USA
    • Corresponding Author InformationCorresponding author. Ross Building, RM 632L, 720 Rutland Avenue, Baltimore, MD 21205, USA. Tel.: +1 410 614 8138; fax: +1 410 502 5862.
    • ,
  • Toby C. Cornish

      Affiliations

    • Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD, USA
    • ,
  • Jie Lu

      Affiliations

    • Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD, USA
    • ,
  • Steve Selvin

      Affiliations

    • School of Public Health, University of California, Berkeley, Berkeley, CA, USA
    • ,
  • Elizabeth Selvin

      Affiliations

    • Departments of Epidemiology and Medicine, Johns Hopkins Bloomberg School of Public Health and the Johns Hopkins School of Medicine, Baltimore, MD, USA

Received 19 August 2008; received in revised form 21 November 2008; accepted 16 December 2008. published online 12 February 2009.

Abstract 

Background

Tissue microarrays (TMAs) are collections of multiple tissue cores placed in parallel in a single acceptor block and traditionally used to investigate protein expression in neoplastic tissues. We validated the use of TMAs to investigate protein expression in vascular segments.

Methods

Vascular tissues were collected from 100 adult subjects undergoing autopsy. A diverse set of vessels were harvested and arrayed over 17 TMAs. A total of 1377 unique tissues, each with a 1.5-mm feature size, were analyzed using histochemical and immunohistochemical (IHC) diaminobenzidine (DAB) methods.

Results

Histomorphometric analysis of vascular disease demonstrated the TMA features captured the majority of the vascular alterations (intimal hyperplasia and atherosclerosis) seen in the original blood vessel section. Measurements of IHC staining intensity based on color deconvolution were used to quantify antigen abundance in defined regions of interest (ROI). Validation was performed using antibodies to connective tissue growth factor (CTGF), receptor for advanced glycation end products (AGER/RAGE), and matrix metalloproteinase 3 (MMP-3). IHC staining was highly correlated between duplicate features from the same vascular site over these three proteins.

Conclusion

This study validates the use of TMA technology to investigate the vascular wall utilizing staining intensity data.

Keywords: Immunohistochemistry, Media, Vascular disease, Human arteries

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 This work was supported by the American Diabetes Association (1-05-JF-20 to MKH) and by the NIH/NIDDK (K01 DK076595 to ES).

PII: S1054-8807(08)00188-9

doi:10.1016/j.carpath.2008.12.007

Cardiovascular Pathology
Volume 19, Issue 3 , Pages 136-146, May 2010

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