Cold-Temperature Plastic Resin Embedding of Liver for DNA- and RNA-Based Genotyping Sydney D. Finkelstein, Rajiv Dhir, Mordechai Rabinovitz, Michelle.

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Cold-Temperature Plastic Resin Embedding of Liver for DNA- and RNA-Based Genotyping Sydney D. Finkelstein, Rajiv Dhir, Mordechai Rabinovitz, Michelle Bischeglia, Patricia A. Swalsky, Petrina DeFlavia, Jeffrey Woods, Anke Bakker, Michael Becich The Journal of Molecular Diagnostics Volume 1, Issue 1, Pages 17-22 (November 1999) DOI: 10.1016/S1525-1578(10)60604-6 Copyright © 1999 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 1 Histological appearance of liver biopsy specimens subject to routine (left, 10% neutral buffered formalin/paraffin embedding) and molecular (right, 2% paraformaldehyde/plastic resin embedding) processing. Liver cells fixed and processed routinely manifest homogeneous velvety cytoplasm with round vesicular nucleus including prominent nucleolus. Liver tissue subject to molecular fixation is indistinguishable in histological appearance. Hematoxylin-eosin; original magnifications, ×250 and ×400. The Journal of Molecular Diagnostics 1999 1, 17-22DOI: (10.1016/S1525-1578(10)60604-6) Copyright © 1999 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 2 Robust amplification of β-actin RNA by RT/PCR (leftmost 8 samples) and epidermal growth factor receptor-2 DNA (rightmost 8 samples) from minute cold-temperature plastic-embedded tissue samples. Microdissected tissue samples consisting of as few as 20 cells from 4-μm-thick histological sections demonstrated strong positive amplification of RNA and DNA gene targets. P, primers only PCR, negative control. The Journal of Molecular Diagnostics 1999 1, 17-22DOI: (10.1016/S1525-1578(10)60604-6) Copyright © 1999 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 3 RT-PCR amplification of hepatitis C virus for amplicons that are short (left, 5′ nontranslated region, 153 bp) and long (right, NS5 region, 364 bp). Left: All samples subject to molecular fixation and processing yielded strong positive amplification without the requirement for nested primer amplification. All samples routinely fixed failed to effectively amplify consistent with total or nearly total loss of viral RNA. Right: A series of HCV-positive and -negative (N) control samples were subject to RT-PCR for specific detection targeting a 364-bp segment of the NS5 region. P, primers only negative control. The Journal of Molecular Diagnostics 1999 1, 17-22DOI: (10.1016/S1525-1578(10)60604-6) Copyright © 1999 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 4 HCV genotyping from plastic resin-embedded tissue section. Attainment of abundant PCR product was found to be the critical factor permitting clear and strong DNA sequencing of tissue HCV. Left:Genotyping based on the 5′ nontranslated region reveals type 1 strains in two samples. Right: NS5 region genotyping was preferred due to greater interstrain sequence variation, which in turn permitted detection and characterization of HCV quasispecies. The Journal of Molecular Diagnostics 1999 1, 17-22DOI: (10.1016/S1525-1578(10)60604-6) Copyright © 1999 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions